FEBS Letters

A change in the sensitivity of elongation factor G to oxidation protects photosystem II from photoinhibition in Synechocystis sp. PCC 6803.

Publication Date: 2012 Jan 28 PMID: 22300643
Authors: Ejima, K. - Kawaharada, T. - Inoue, S. - Kojima, K. - Nishiyama, Y.
Journal: FEBS Lett

The repair of photosystem II (PSII) after photodamage is particularly sensitive to oxidative stress and inhibition of such repair is associated with the oxidation of specific cysteine residues in elongation factor G (EF-G), a key translation factor, in the cyanobacterium Synechocystis sp. PCC 6803. Expression of mutated EF-G with a target cysteine residue replaced by serine in Synechocystis resulted in the protection of PSII from photoinhibition. This protection was attributable to the enhanced repair of PSII via acceleration of the synthesis of the D1 protein, which might have been due to reduced sensitivity of protein synthesis to oxidative stress.

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Kaiso uses all three zinc fingers and adjacent sequence motifs for high affinity binding to sequence-specific and methyl-CpG DNA targets.

Publication Date: 2012 Jan 30 PMID: 22300642
Authors: Buck-Koehntop, B. A. - Martinez-Yamout, M. A. - Jane Dyson, H. - Wright, P. E.
Journal: FEBS Lett

Kaiso is a Cys(2)His(2) zinc finger (ZF) protein that mediates methyl-CpG-dependent and sequence-specific transcriptional repression. As a first step towards elucidating the structural and molecular basis for recognition of these disparate DNA sequences, the minimal binding region of Kaiso was identified and optimal DNA sequences for high-affinity interactions were characterized. Contrary to previous findings, Kaiso requires all three zinc fingers plus adjacent protein regions for DNA recognition. An N-terminal extension contributes to structural stability, while an extended C-terminal region augments DNA binding. Complexes formed between the optimized Kaiso construct and both DNA sequences are suitable for future structural evaluation.

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HBXIP upregulates CD46, CD55 and CD59 through ERK1/2/NF-kappaB signaling to protect breast cancer cells from complement attack.

Publication Date: 2012 Jan 27 PMID: 22293503
Authors: Cui, W. - Zhao, Y. - Shan, C. - Kong, G. - Hu, N. - Zhang, Y. - Zhang, S. - Zhang, W. - Zhang, Y. - Zhang, X. - Ye, L.
Journal: FEBS Lett

Hepatitis B X-interacting protein (HBXIP) is able to enhance migration of breast cancer cells. However, the role of HBXIP in regulation of complement-dependent cytotoxicity (CDC) in breast cancer is not understood. Here, we report that HBXIP contributes to protecting breast cancer cells from CDC by upregulating membrane-bound complement regulatory protein (mCRPs), including CD46, CD55 and CD59. We found that HBXIP upregulated mCRPs through activating p-ERK1/2/NF-kappaB. Interestingly, the knockdown of CD59 was able to block the HBXIP-enhanced breast tumor growth in animal. Thus, we conclude that HBXIP upregulates CD46, CD55 and CD59 through p-ERK1/2/NF-kappaB signaling to protect breast cancer from CDC.

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Selenomodification of tRNA in archaea requires a bipartite rhodanese enzyme.

Publication Date: 2012 Jan 27 PMID: 22293502
Authors: Su, D. - Ojo, T. T. - Soll, D. - Hohn, M. J.
Journal: FEBS Lett

5-Methylaminomethyl-2-selenouridine (mnm(5)Se(2)U) is found in the first position of the anticodon in certain tRNAs from bacteria, archaea and eukaryotes. This selenonucleoside is formed in Escherichia coli from the corresponding thionucleoside mnm(5)S(2)U by the monomeric enzyme YbbB. This nucleoside is present in the tRNA of Methanococcales, yet the corresponding 2-selenouridine synthase is unknown in archaea and eukaryotes. Here we report that a bipartite ybbB ortholog is present in all members of the Methanococcales. Gene deletions in Methanococcus maripaludis and in vitro activity assays confirm that the two proteins act in trans to form in tRNA a selenonucleoside, presumably mnm(5)Se(2)U. Phylogenetic data suggest a primal origin of seleno-modified tRNAs.

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Oligomeric interactions of TGF-beta and BMP receptors.

Publication Date: 2012 Jan 27 PMID: 22293501
Authors: Ehrlich, M. - Gutman, O. - Knaus, P. - Henis, Y. I.
Journal: FEBS Lett

Transforming growth factor-beta (TGF-beta) and bone morphogenetic protein (BMP) cytokines participate in a multiplicity of ways in the regulation of numerous physiological and pathological processes. Their wide-ranging biological functions are controlled by several mechanisms, including regulation of transcription, complex formation among the signaling receptors (oligomerization) and with co-receptors, binding of the receptors to scaffolding proteins or their targeting to specific membrane domains. Here, we address the generation of TGF-beta and BMP receptor homo- and hetero-oligomers and its roles as a mechanism capable of fast regulation of signaling by these crucial cytokines. We examine the available biochemical, biophysical and structural evidence for the ternary structure of these complexes, and the possible roles of homomeric and heteromeric receptor oligomers in signaling.

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MIG-13 controls anteroposterior cell migration by interacting with UNC-71/ADM-1 and SRC-1 in Caenorhabditis elegans.

Publication Date: 2012 Jan 27 PMID: 22293500
Authors: Masuda, H. - Nakamura, K. - Takata, N. - Itoh, B. - Hirose, T. - Moribe, H. - Mekada, E. - Okada, M.
Journal: FEBS Lett

The transmembrane protein MIG-13 is a key regulator required for anterior migration of neural cells in Caenorhabditis elegans, but the signaling mechanisms involved remain unknown. Here, we isolated a suppressor mutation in the unc-71/adm-1 gene, which rescued the AVM neuron migration defect in mig-13 mutants. Genetic analyses revealed that UNC-71 at least partly acts downstream of MIG-13 and has an inhibitory effect on the anterior cell migration. The unc-71 mutation also rescued the anterior migration defect of AVM neuron in src-1 mutants. These findings suggest that MIG-13 controls anteroposterior cell migration by interacting with UNC-71 and SRC-1 in C. elegans.

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Overexpression of lalA, a paralog of labA, is capable of affecting both circadian gene expression and cell growth in the cyanobacterium Synechococcus elongatus PCC 7942.

Publication Date: 2012 Jan 27 PMID: 22289183
Authors: Taniguchi, Y. - Nishikawa, T. - Kondo, T. - Oyama, T.
Journal: FEBS Lett

In the cyanobacterium Synechococcus elongatus, LabA negatively regulates circadian gene expression under the control of Kai-protein-based clock. Here we conducted a molecular genetic analysis of lalA, a paralog of labA. Although a lalA loss of function mutant did not exhibit any apparent phenotype under our experimental conditions, lalA overexpression inhibited cell growth and decreased cell viability. Moderate lalA overexpression brought about abnormalities in circadian gene expression: reduced amplitude of kaiBC expression rhythm, and altered peak and trough timing of psbAI and kaiA expression rhythms. These results imply that lalA is capable of affecting circadian gene expression and cell growth.

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Saccharomyces cerevisiae gene YMR291W/TDA1 mediates the in vivo phosphorylation of hexokinase isoenzyme 2 at serine-15.

Publication Date: 2012 Jan 27 PMID: 22289182
Authors: Kettner, K. - Krause, U. - Mosler, S. - Bodenstein, C. - Kriegel, T. - Rodel, G.
Journal: FEBS Lett

Hxk2 is the predominant hexokinase of Saccharomyces cerevisiae during growth on glucose. In addition to its role in glycolysis, the enzyme is involved in glucose sensing and regulation of gene expression. Glucose limitation causes the phosphorylation of Hxk2 at serine-15 which affects the nucleo-cytoplasmic distribution and dimer stability of the enzyme. In order to identify the responsible kinase, we screened selected protein kinase single-gene deletion mutants by high resolution clear native PAGE. Deletion of YMR291W/TDA1 resulted in the absence of the Hxk2 phosphomonomer, indicating an indispensable role of the corresponding protein in Hxk2 phosphorylation.

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N-Acetyl lysyl-tRNA synthetases evolved by a CcdB-based selection possess N-acetyl lysine specificity in vitro and in vivo.

Publication Date: 2012 Jan 27 PMID: 22289181
Authors: Umehara, T. - Kim, J. - Lee, S. - Guo, L. T. - Soll, D. - Park, H. S.
Journal: FEBS Lett

Posttranslational modifications play a crucial role in modulating protein structure and function. Genetic incorporation of unnatural amino acids into a specific site of a protein facilitates the systematic study of protein modifications including acetylation. We here report the directed evolution of pyrrolysyl-tRNA synthetase (PylRS) from Methanosarcina mazei to create N-acetyl lysyl-tRNA synthetases (AcKRSs) using a new selection system based on the killing activity of the toxic ccdB gene product. The amino acid specificity of these and of published [1,2] AckRSs was tested in vitro and in vivo, and the enzyme-kinetic properties of the AckRSs were evaluated for the first time.

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Crystal structure of cce_0566 from Cyanothece 51142, a protein associated with nitrogen fixation in the DUF269 family.

Publication Date: 2012 Jan 27 PMID: 22289180
Authors: Buchko, G. W. - Robinson, H.
Journal: FEBS Lett

The crystal structure for cce_0566 (171 aa, 19.4kDa), a DUF269 annotated protein from the diazotrophic cyanobacterium Cyanothece sp. ATCC 51142, was determined to 1.60A resolution. cce_0566 is a homodimer with each molecule composed of eight a-helices folded on one side of a three strand anti-parallel beta-sheet. Hydrophobic interactions between the side chains of largely conserved residues on the surface of each beta-sheet hold the dimer together. The fold observed for cce_0566 may be unique to proteins in the DUF269 family, hence, the protein may also have a function unique to nitrogen fixation. A solvent accessible cleft containing conserved charged residues near the dimer interface could represent the active site or ligand-binding surface for the protein's biological function. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: DUF269 and DUF269bind by x-ray crystallography (View interaction).

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Transcription-independent role of Bach1 in mitosis through a nuclear exporter Crm1-dependent mechanism.

Publication Date: 2012 Jan 27 PMID: 22289179
Authors: Li, J. - Shiraki, T. - Igarashi, K.
Journal: FEBS Lett

The transcriptional repressor Bach1 mediates various stress responses. Despite its role in transcription, Bach1 is predominantly exported to the cytoplasm in a Crm1-dependent manner, but the functional role of its cytoplasmic retention is still unclear. We found that Bach1 was also excluded from mitotic chromatin by a C-terminal cytoplasmic localization sequence dependent and leptomycin B sensitive process. Bach1 depletion resulted in disordered mitotic chromosome alignment, which was rescued by Bach1 mutants lacking the BTB or DNA binding domains, suggesting its transcription-independent mechanism. We thus revealed a novel role of Bach1 in the regulation of mitotic chromosome dynamics.

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Cataract-linked gammaD-crystallin mutants have weak affinity to lens chaperones alpha-crystallins.

Publication Date: 2012 Jan 27 PMID: 22289178
Authors: Mishra, S. - Stein, R. A. - McHaourab, H. S.
Journal: FEBS Lett

To test the hypothesis that alpha-crystallin chaperone activity plays a central role in maintenance of lens transparency, we investigated its interactions with gamma-crystallin mutants that cause congenital cataract in mouse models. Although the two substitutions, I4F and V76D, stabilize a partially unfolded gammaD-crystallin intermediate, their affinities to alpha-crystallin are marginal even at relatively high concentrations. Detectable binding required further reduction of gammaD-crystallin stability which was achieved by combining the two mutations. Our results demonstrate that mutants and possibly age-damaged gamma-crystallin can escape quality control by lens chaperones rationalizing the observation that they nucleate protein aggregation and lead to cataract. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: gammaD Crystallin and alphaB Crystallinbind by molecular sieving (View interaction) gammaD Crystallin and alphaB Crystallinbind by fluorescence technology (View interaction) gammaD Crystallin and alphaA Crystallinbind by fluorescence technology (View interaction) gammaD Crystallin and alphaA Crystallinbind by molecular sieving (View interaction).

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Delivery of Bordetella pertussis adenylate cyclase toxin to target cells via outer membrane vesicles.

Publication Date: 2012 Jan 27 PMID: 22289177
Authors: Donato, G. M. - Goldsmith, C. S. - Paddock, C. D. - Eby, J. C. - Gray, M. C. - Hewlett, E. L.
Journal: FEBS Lett

Bordetella pertussis adenylate cyclase toxin (ACT) intoxicates cells by producing intracellular cAMP. B. pertussis outer membrane vesicles (OMV) contain ACT on their surface (OMV-ACT), but the properties of OMV-ACT were previously unknown. We found that B. pertussis in the lung from a fatal pertussis case contains OMV, suggesting an involvement in pathogenesis. OMV-ACT and ACT intoxicate cells with and without the toxin's receptor CD11b/CD18. Intoxication by ACT is blocked by antitoxin and anti-CD11b antibodies, but not by cytochalasin-D; in contrast, OMV-ACT is unaffected by either antibody and blocked by cytochalasin-D. Thus OMV-ACT can deliver ACT by processes distinct from those of ACT alone.

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Cyclin-dependent kinase 4 is a novel target in micoRNA-195-mediated cell cycle arrest in bladder cancer cells.

Publication Date: 2012 Jan 27 PMID: 22289176
Authors: Lin, Y. - Wu, J. - Chen, H. - Mao, Y. - Liu, Y. - Mao, Q. - Yang, K. - Zheng, X. - Xie, L.
Journal: FEBS Lett

miRNAs are a class of small-noncoding RNAs capable of negatively regulating gene expression. Here, we found that miR-195 is down-regulated in human bladder cancer tissue versus normal adjacent tissue. To better characterize the role of miR-195 in bladder cancer, we conducted gain of function analysis by transfecting bladder cancer cell line T24 with chemically synthesized miR-195 mimic. We identified CDK4, an early G1 cell cycle regulator, as a novel target of miR-195. Selective over-expression of miR-195 could induce G1-phase arrest in T24 cells, and subsequently inhibit T24 cell growth. These findings indicate that miR-195 could be a potential tumor suppressor in bladder cancer.

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Novel assay with fluorescence-labelled PrP peptides for differentiating L-type atypical and classical BSEs, and scrapie.

Publication Date: 2012 Jan 25 PMID: 22285492
Authors: Kasai, K. - Hitara, A. - Ohyama, T. - Nokihara, K. - Yokoyama, T. - Mohri, S.
Journal: FEBS Lett

Characteristic differences of prions may account for the conformational diversity of the pathogenic isoform of prion protein (PrP(Sc)). Here, we applied a protein detection procedure by using fluorescent-labelled peptides for detecting PrP(Sc). Five prion protein (PrP) related peptides were found to change significantly their fluorescent intensities with prion-affected animal samples. Their reactivity was different among atypical L-BSE, classical BSE and scrapie. The pull-down assay revealed that they precipitated PrP(Sc) specifically. These findings suggest that fluorescent intensity changes depend on peptide-PrP(Sc) binding. This novel approach may distinguish the fine structural differences in PrP(Sc), which were not detected by the pull-down assay. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS:: PrP-peptide HPP01-03physically interacts with sheep PrP by pull down (View Interaction) PrP-peptide HPP06physically interacts with sheep PrP by pull down (View Interaction) PrP-peptide HPP11physically interacts with sheep PrP by pull down (View Interaction) PrP-peptide HPP06physically interacts with mouse PrP by pull down (View Interaction) PrP-peptide HPP11physically interacts with mouse PrP by pull down (View Interaction) PrP-peptide HPP01-03physically interacts with mouse PrP by pull down (View Interaction) PrP-peptide HPP01-03physically interacts with cattle PrP by pull down (View Interaction) PrP-peptide HPP06physically interacts with cattle PrP by pull down (View Interaction) PrP-peptide HPP11physically interacts with cattle PrP by pull down (View Interaction).

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RUVBL2 is a novel repressor of ARF transcription.

Publication Date: 2012 Jan 25 PMID: 22285491
Authors: Xie, C. - Wang, W. - Yang, F. - Wu, M. - Mei, Y.
Journal: FEBS Lett

ARF is the second most commonly inactivated tumor suppressor behind p53. It has been implicated in the control of cell proliferation, cell senescence, and tumor suppression. However, the detailed mechanism underlying the transcriptional control of ARF remains largely unknown. Here we report RUVBL2 as a novel transcriptional repressor of ARF. Ectopic expression of RUVBL2 decreases the levels of ARF, whereas knockdown of RUVBL2 results in a marked increase in ARF levels. In addition, RUVBL2 down-regulates the levels of p53 in an ARF-dependent manner. Mechanistically, RUVBL2 binds to the distal region of ARF promoter, thus leading to the repression of ARF transcription. These results suggest an important role of RUVBL2 in the regulation of ARF-p53 pathway.

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LY294002 inhibits TLR3/4-mediated IFN-beta production via inhibition of IRF3 activation with a PI3K-independent mechanism.

Publication Date: 2012 Jan 25 PMID: 22285490
Authors: Zhao, W. - Qi, J. - Wang, L. - Zhang, M. - Wang, P. - Gao, C.
Journal: FEBS Lett

TLR3 and TLR4 utilize adaptor TRIF to activate interferon regulatory factor 3 (IRF3), resulting in interferon beta (IFN-beta) production to mediate anti-viral infection. In this report, we analyzed the effect of two known phosphatidylinositol 3-kinase (PI3K) inhibitors LY294002 and wortmannin on LPS- and poly(I:C)-induced IFN-beta production in peritoneal macrophages. LY294002 inhibited LPS- and poly(I:C)-induced IFN-beta transcription and secretion. In contrast, wortmannin could not inhibit IFN-beta production. Furthermore, IRF3 transcriptional activation and binding to IFN-beta promoter were found to be inhibited by LY294002. Therefore, our findings demonstrate LY294002 negatively regulates LPS- and poly(I:C)-induced IFN-beta production through inhibition of IRF3 activation in a PI3K-independent manner.

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Phosphate homeostasis in the yeast Saccharomyces cerevisiae, the key role of the SPX domain-containing proteins.

Publication Date: 2012 Jan 25 PMID: 22285489
Authors: Secco, D. - Wang, C. - Shou, H. - Whelan, J.
Journal: FEBS Lett

In the yeast Saccharomyces cerevisiae, a working model for nutrient homeostasis in eukaryotes, inorganic phosphate (Pi) homeostasis is regulated by the PHO pathway, a set of phosphate starvation induced genes, acting to optimize Pi uptake and utilization. Among these, a subset of proteins containing the SPX domain has been shown to be key regulators of Pi homeostasis. In this review, we summarize the recent progresses in elucidating the mechanisms controlling Pi homeostasis in yeast, focusing on the key roles of the SPX domain-containing proteins in these processes, as well as describing the future challenges and opportunities in this fast-moving field.

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Scythe cleavage during Fas (APO-1)-and staurosporine-mediated apoptosis.

Publication Date: 2012 Jan 25 PMID: 22285488
Authors: Preta, G. - Fadeel, B.
Journal: FEBS Lett

Scythe is a nuclear protein that has been implicated in the apoptotic process in Drosophila melanogaster; however, its role in apoptosis of mammalian cells is not fully elucidated. Here we show that cleavage of Scythe by caspase-3 occurs after activation of both the extrinsic (i.e. Fas/APO-1-mediated) and the intrinsic (i.e. staurosporine-induced) apoptosis pathway. Moreover, this caspase-dependent cleavage correlates with Scythe translocation from the nucleus to the cytosol. We also show that cytosolic re-localization of Scythe is required for Fas/APO-1-triggered phosphatidylserine (PS) exposure, a signal for macrophage clearance of apoptotic cells. Our data suggest that Scythe cleavage may represent a marker for caspase-3 activation and implicate cytosolic re-localization of Scythe in the pathway of PS exposure.

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The beta1 domain of protein G can replace the chorismate mutase domain of the T-protein.

Publication Date: 2012 Jan 25 PMID: 22285487
Authors: Osuna, J. - Flores, H. - Saab-Rincon, G.
Journal: FEBS Lett

T-protein is composed of chorismate mutase (AroQ(T)) fused to the N-terminus of prephenate dehydrogenase (TyrA). Here, we report the replacement of AroQ(T) with the beta1-domain of protein G (Gbeta1). The TyrA domain shows a strong dehydrogenase activity within the context of this fusion, and our data indicate that Gbeta1-TyrA folds into a dimeric conformation. Amino acid substitutions in the Gbeta1 domain of Gbeta1-TyrA identified residues involved in stabilizing the TyrA dimeric conformation. Gbeta1 substitutions in the N-terminal beta-hairpin eliminated Gbeta1-TyrA expression, whereas Gbeta1-TyrA tolerated Gbeta1 substitutions in the C-terminal beta-hairpin and in the alpha-helix. All of the characterized variants folded into a dimeric conformation. The importance of the beta2-strand in forming a Gbeta1 homo-dimerization interface explains the relevance of the first-beta-hairpin in stabilizing the dimeric TyrA protein.

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MIZ1, an essential protein for root hydrotropism, is associated with the cytoplasmic face of the endoplasmic reticulum membrane in Arabidopsis root cells.

Publication Date: 2012 Jan 24 PMID: 22285304
Authors: Yamazaki, T. - Miyazawa, Y. - Kobayashi, A. - Moriwaki, T. - Fujii, N. - Takahashi, H.
Journal: FEBS Lett

MIZ1 is encoded by a gene essential for root hydrotropism in Arabidopsis. To characterize the property of MIZ1, we used transgenic plants expressing GFP-tagged MIZ1 (MIZ1-GFP) and mutant MIZ1 (MIZ1(G235E)-GFP) in a miz1-1 mutant. Although both chimeric genes were transcribed, the translational products of MIZ1(G235E)-GFP did not accumulate in roots. Moreover, MIZ1-GFP complemented the mutant phenotype but not MIZ1(G235E)-GFP. The signal corresponding to MIZ1-GFP was detected at high levels in cortical cells and lateral root cap cells and accumulated in compartments in cortical cells. MIZ1-GFP was fractionated into a soluble protein fraction and an endoplasmic reticulum (ER) membrane fraction, where it was bound to the surface of the ER membrane at the cytosolic side.

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Is the chromanol head group of vitamin E nature's final truth on chain-breaking antioxidants?

Publication Date: 2012 Jan 23 PMID: 22281199
Authors: Ohlow, M. J. - Granold, M. - Schreckenberger, M. - Moosmann, B.
Journal: FEBS Lett

Tocopherol is believed to be the most potent naturally occurring chain-breaking antioxidant. Hence, its refined phenolic head group chromanol may represent an optimum evolutionary solution to the problem of free-radical chain reactions in the lipid bilayer. To test the universal validity of this assumption beyond phenolic head groups, we have synthesized aromatic amine analogues of vitamin E and trolox with otherwise closely matching physicochemical properties: NH-toc and NH-trox. We have found that NH-toc and NH-trox were significantly more potent free radical scavengers, lipid peroxidation inhibitors and cytoprotective agents than their phenolic templates, tocopherol and trolox. In a chemical sense, thus, the chromanol head group does not constitute a global optimum for the design of chain-breaking antioxidants.

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Mitochondrial ATP-sensitive potassium channel activity and hypoxic preconditioning are independent of an inwardly rectifying potassium channel subunit in Caenorhabditis elegans.

Publication Date: 2012 Jan 21 PMID: 22281198
Authors: Wojtovich, A. P. - Distefano, P. - Sherman, T. - Brookes, P. S. - Nehrke, K.
Journal: FEBS Lett

Hypoxic preconditioning (HP) is an evolutionarily-conserved mechanism that protects an organism against stress. The mitochondrial ATP-sensitive K(+) channel (mK(ATP)) plays an essential role in the protective signaling, but remains molecularly undefined. Several lines of evidence suggest that mK(ATP) may arise from an inward rectifying K(+) channel (Kir). The genetic model organism Caenorhabditis elegans exhibits HP and displays mK(ATP) activity. Here, we investigate the tissue expression profile of the three C. elegans Kir genes and demonstrate that mutant strains where the irk genes have been deleted either individually or in combination can be protected by HP and exhibit robust mK(ATP) channel activity in purified mitochondria. These data suggest that the mK(ATP) in C. elegans does not arise from a Kir derived channel.

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Dimerization, but not phosphothreonine binding, is conserved between the forkhead-associated domains of Drosophila MU2 and human MDC1.

Publication Date: 2012 Jan 21 PMID: 22273583
Authors: Luo, S. - Ye, K.
Journal: FEBS Lett

Mutator 2 (MU2) in Drosophila melanogaster has been proposed to be the ortholog of human MDC1, a key mediator in DNA damage response. The forkhead-associated (FHA) domain of MDC1 is a dimerization module regulated by trans binding to phosphothreonine 4 from another molecule. Here we present the crystal structure of the MU2 FHA domain at 1.9A resolution, revealing its evolutionarily conserved role in dimerization. As compared to the MDC1 FHA domain, the MU2 FHA domain dimerizes using a different and more stable interface and contains a degenerated phosphothreonine-binding pocket. Our results suggest that the MU2 dimerization is constitutive and lacks phosphorylation-mediated regulation. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: MU2 and MU2bind by cosedimentation in solution (View interaction) MU2 and MU2bind by X-ray crystallography (View interaction) MU2 and MU2bind by molecular sieving (View interaction).

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Ribosome-independent biosynthesis of biologically active peptides: Application of synthetic biology to generate structural diversity.

Publication Date: 2012 Jan 21 PMID: 22273582
Authors: Giessen, T. W. - Marahiel, M. A.
Journal: FEBS Lett

Peptide natural products continue to play an important role in modern medicine as last-resort treatments of many life-threatening diseases, as they display many interesting biological activities ranging from antibiotic to antineoplastic. A large fraction of these microbial natural products is assembled by ribosome-independent mechanisms. Progress in sequencing technology and the mechanistic understanding of secondary metabolite pathways has led to the discovery of many formerly cryptic natural products and a molecular understanding of their assembly. Those advances enable us to apply protein and metabolic engineering approaches towards the manipulation of biosynthetic pathways. In this review we discuss the application potential of both templated and non-templated pathways as well as chemoenzymatic strategies for the structural diversification and tailoring of peptide natural products.

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Unusual NADPH conformation in the crystal structure of a cinnamyl alcohol dehydrogenase from Helicobacter pylori in complex with NADP(H) and substrate docking analysis.

Publication Date: 2012 Jan 21 PMID: 22269576
Authors: Seo, K. H. - Zhuang, N. - Chen, C. - Song, J. Y. - Kang, H. L. - Rhee, K. H. - Lee, K. H.
Journal: FEBS Lett

Cinnamyl alcohol dehydrogenase is a zinc- and NADPH-dependent dehydrogenase catalyzing the reversible conversion of p-hydroxycinnamaldehydes to their corresponding hydroxycinnamyl alcohols. A CAD homolog from Helicobacter pylori (HpCAD) possesses broad substrate specificities like the plant CADs and additionally a dismutation activity converting benzaldehyde to benzyl alcohol and benzoic acid. We have determined the crystal structure of HpCAD complexed with NADP(H) at 2.18A resolution to get a better understanding of this class of CAD outside of plants. The structure of HpCAD is highly homologous to the sinapyl alcohol dehydrogenase and the plant CAD with well-conserved residues involved in catalysis and zinc binding. However, the NADP(H) binding mode of the HpCAD has been found to be significantly different from those of plant CADs. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: HpCAD and HpCADbind by x-ray crystallography (View interaction).

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Human synaptotagmin-II is not a high affinity receptor for botulinum neurotoxin B and G: Increased therapeutic dosage and immunogenicity.

Publication Date: 2012 Jan 16 PMID: 22265973
Authors: Strotmeier, J. - Willjes, G. - Binz, T. - Rummel, A.
Journal: FEBS Lett

Botulinum neurotoxins (BoNTs) inhibit neurotransmitter release by hydrolysing SNARE proteins essential for exocytosis. The synaptic vesicle protein synaptotagmin-II of rat and mouse acts as neuronal high affinity receptor for BoNT/B and BoNT/G. Here, we show that human synaptotagmin-II is not a high affinity receptor for BoNT/B and G due to a phenylalanine to leucine mutation in its luminal domain present only in humans and chimpanzees. It eliminates one of three major interactions between synaptotagmin-II and BoNT/B and hereby explains the disparity in potency of BoNT/B in humans and mice as well as the 40-fold higher dosage of rimabotulinumtoxinB versus onabotulinumtoxinA. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: rSyt-IIbinds to BoNT/G by pull down (View Interaction: 1, 2) rSyt-IIbinds to BoNT/B by pull down (View Interaction: 1, 2).

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Crystal structures of the coil 2B fragment and the globular tail domain of human lamin B1.

Publication Date: 2012 Jan 16 PMID: 22265972
Authors: Ruan, J. - Xu, C. - Bian, C. - Lam, R. - Wang, J. P. - Kania, J. - Min, J. - Zang, J.
Journal: FEBS Lett

We present here the crystal structures of human lamin B1 globular tail domain and coiled 2B domain, which adopt similar folds to Ig-like domain and coiled-coil domain of lamin A, respectively. Despite the overall similarity, we found an extra intermolecular disulfide bond in the lamin B1 coil 2B domain, which does not exist in lamin A/C. In addition, the structural analysis indicates that interactions at the lamin B1 homodimer interface are quite different from those of lamin A/C. Thus our research not only reveals the diversely formed homodimers among lamin family members, but also sheds light on understanding the important roles of lamin B1 in forming the nuclear lamina matrix. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: Lamin-B and Lamin-Bbind by x-ray crystallography (View interaction).

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MicroRNA-195-5p suppresses glucose uptake and proliferation of human bladder cancer T24 cells by regulating GLUT3 expression.

Publication Date: 2012 Jan 16 PMID: 22265971
Authors: Fei, X. - Qi, M. - Wu, B. - Song, Y. - Wang, Y. - Li, T.
Journal: FEBS Lett

It has been reported that expression of glucose transporter member 3 (GLUT3) is up-regulated in bladder cancers. However, the regulating mechanism remains unknown. Here, we assessed whether microRNAs (miRNAs) regulate GLUT3 expression in bladder cancers. In our study, miR-195-5p was identified to directly targeted GLUT3 3'-untranslated region (UTR) in bladder cancer T24 cells. Small interfering RNA (siRNA)- and miR-195-5p-mediated GLUT3 knockdown experiments revealed that miR-195-5p decreased T24 cells glucose uptake, inhibited cell growth and promoted cell apoptosis through suppression of GLUT3 expression. Therefore, miR-195-5p is a novel and also the first identified miRNA that targets GLUT3, and the aberrant decreased expression of miR-195-5p and consequent GLUT3 up-regulation may contribute to bladder carcinogenesis.

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Phosphosite conservation in single domain orthologs versus paralogs: A way to combine differential regulation with redundant core functions.

Publication Date: 2012 Jan 18 PMID: 22265693
Authors: Huyck, L. - Van Troys, M. - Ampe, C.
Journal: FEBS Lett

Evolutionary conservation for structure function relations is commonly accepted. Here we hypothesize that closely related single domain paralogous proteins, having similar expression profiles and redundant biochemical core functions, additionally evolved to allow and maintain isoform specific differential regulation by single conserved amino acid substitutions. To substantiate this, we considered two families of closely related actin binding proteins combined with data mining of phosphorylated residues in human and mouse proteins. We show that such residues are identical in other orthologs whereas paralogs have a different, but also conserved, non-phosphorylatable residue at the equivalent positions.

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Follistatin-related protein/follistatin-like 1 evokes an innate immune response via CD14 and toll-like receptor 4.

Publication Date: 2012 Jan 17 PMID: 22265692
Authors: Murakami, K. - Tanaka, M. - Usui, T. - Kawabata, D. - Shiomi, A. - Iguchi-Hashimoto, M. - Shimizu, M. - Yukawa, N. - Yoshifuji, H. - Nojima, T. - Ohmura, K. - Fujii, T. - Umehara, H. - Mimori, T.
Journal: FEBS Lett

Follistatin-related protein (FRP)/follistatin-like 1 (FSTL1) has multi-specific binding nature especially with TGF-beta superfamily proteins, and thereby modulates organ development. However, its function in immune systems remains unclear. Previously, we reported FRP interacts with CD14, which is known to mediate toll-like receptor 4 (TLR4) signaling. Here, we investigated whether FRP activates TLR4 signaling. Recombinant FRP induced interleukin 6 or interleukin 8 production from target cells in a CD14- and TLR4-dependent manner. Moreover, similar to lipopolysaccharide (LPS), FRP induced tolerance to the second LPS stimulation. FRP has the function of evoking innate immune responses as one of the endogenous TLR4 agonists. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS:: FRPphysically interacts with CD14 by fluorescence-activated cell sorting (View interaction).

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MiR-18a regulates expression of the pancreatic transcription factor Ptf1a in pancreatic progenitor and acinar cells.

Publication Date: 2012 Jan 17 PMID: 22265691
Authors: Yang, Y. - Ding, L. - An, Y. - Zhang, Z. W. - Lang, Y. - Tai, S. - Guo, F. - Teng, C. B.
Journal: FEBS Lett

The basic helix-loop-helix (bHLH) transcription factor Ptf1a plays stage-specific roles in the developing pancreas. During early pancreatic development, low levels of Ptf1a preferentially promote the differentiation of pancreatic progenitor cells into endocrine cells, whereas high levels of Ptf1a shift pancreatic progenitors towards an exocrine cell fate. In adults, Ptf1a is essential for the production of exocrine enzymes by pancreatic acinar cells. In this paper, we show that Ptf1a expression is repressed by miR-18a in pancreatic progenitors and acinar cells via its binding to the 3'UTR of Ptf1a mRNA. Furthermore, overexpression of miR-18a exerts little effect on pancreatic progenitors and acinar cells. These results indicate that miR-18a plays a fine-tuning role in regulating pancreatic progenitors and exocrine cells through the repression of Ptf1a expression.

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Verification of the interdomain contact site in the inactive monomer, and the domain-swapped fold in the active dimer of Hsp33 in solution.

Publication Date: 2012 Jan 17 PMID: 22265690
Authors: Lee, Y. S. - Ryu, K. S. - Kim, S. J. - Ko, H. S. - Sim, D. W. - Jeon, Y. H. - Kim, E. H. - Choi, W. S. - Won, H. S.
Journal: FEBS Lett

Upon dimerization by oxidation, Hsp33 functions as a molecular chaperone in prokaryotes. Previously published structures of both the inactive and active species are of doubtful relevance to the solution conformations since the inactive (reduced) crystal structure was dimeric, while the active (oxidized) species was crystallized with a truncation of its regulation domain. The interdomain contact site of the inactive monomer, identified in this work, is consistent with that previously observed in the reduced dimer crystal. In contrast, fluorescence quenching of the active dimer contradicted the results expected from the domain-swapped fold observed in the truncated dimer crystal. The results of this study provide important new information concerning controversial issues in the activation process of Hsp33.

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Approaches to chemical synthetic biology.

Publication Date: 2012 Jan 17 PMID: 22265689
Authors: Chiarabelli, C. - Stano, P. - Anella, F. - Carrara, P. - Luisi, P. L.
Journal: FEBS Lett

Synthetic biology is first represented in terms of two complementary aspects, the bio-engineering one, based on the genetic manipulation of extant microbial forms in order to obtain forms of life which do not exist in nature; and the chemical synthetic biology, an approach mostly based on chemical manipulation for the laboratory synthesis of biological structures that do not exist in nature. The paper is mostly devoted to shortly review chemical synthetic biology projects currently carried out in our laboratory. In particular, we describe: the minimal cell project, then the "Never Born Proteins" and lastly the Never Born RNAs. We describe and critically analyze the main results, emphasizing the possible relevance of chemical synthetic biology for the progress in basic science and biotechnology.

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Single-particle tracking of immunoglobulin E receptors (FcepsilonRI) in micron-sized clusters and receptor patches.

Publication Date: 2012 Jan 18 PMID: 22265688
Authors: Spendier, K. - Lidke, K. A. - Lidke, D. S. - Thomas, J. L.
Journal: FEBS Lett

When mast cells contact a monovalent antigen-bearing fluid lipid bilayer, IgE-loaded FcepsilonRI receptors aggregate at contact points and trigger degranulation and the release of immune activators. We used two-color total internal reflection fluorescence microscopy and single-particle tracking to show that most fluorescently labeled receptor complexes diffuse freely within these micron-size clusters, with a diffusion coefficient comparable to free receptors in resting cells. At later times, when the small clusters coalesce to form larger patches, receptors diffuse even more rapidly. In all cases, Monte Carlo diffusion simulations ensured that the tracking results were free of bias, and distinguished biological from statistical variation. These results show the diversity in receptor mobility in mast cells, demonstrating at least three distinct states of receptor diffusivity.

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Back-reactions, short-circuits, leaks and other energy wasteful reactions in biological electron transfer: Redox tuning to survive life in O(2).

Publication Date: 2012 Jan 13 PMID: 22251618
Authors: Rutherford, A. W. - Osyczka, A. - Rappaport, F.
Journal: FEBS Lett

The energy-converting redox enzymes perform productive reactions efficiently despite the involvement of high energy intermediates in their catalytic cycles. This is achieved by kinetic control: with forward reactions being faster than competing, energy-wasteful reactions. This requires appropriate cofactor spacing, driving forces and reorganizational energies. These features evolved in ancestral enzymes in a low O(2) environment. When O(2) appeared, energy-converting enzymes had to deal with its troublesome chemistry. Various protective mechanisms duly evolved that are not directly related to the enzymes' principal redox roles. These protective mechanisms involve fine-tuning of reduction potentials, switching of pathways and the use of short circuits, back-reactions and side-paths, all of which compromise efficiency. This energetic loss is worth it since it minimises damage from reactive derivatives of O(2) and thus gives the organism a better chance of survival. We examine photosynthetic reaction centres, bc(1) and b(6)f complexes from this view point. In particular, the evolution of the heterodimeric PSI from its homodimeric ancestors is explained as providing a protective back-reaction pathway. This "sacrifice-of-efficiency-for-protection" concept should be generally applicable to bioenergetic enzymes in aerobic environments.

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Identification of essential sequences for cellular localization in the muscle-specific ubiquitin E3 ligase MAFbx/Atrogin 1.

Publication Date: 2012 Jan 11 PMID: 22249105
Authors: Julie, L. C. - Sabrina, B. P. - Marie-Pierre, L. - Leibovitch, S. A.
Journal: FEBS Lett

In skeletal muscle atrophy, upregulation and nuclear accumulation of the Ubiquitin E3 ligase MAFbx is essential for accelerated muscle protein loss, but the nuclear/cytoplasmic shuttling of MAFbx is undefined. Here we found that MAFbx contains two functional nuclear localization signals (NLS). Mutation or deletion of only one NLS induced cytoplasmic localization of MAFbx. We identified a non-classical NES located in the leucine charged domain (LCD) of MAFbx, which is leptomycin B insensitive. We demonstrated that mutation (L169Q) in LLXXL motif of LCD suppressed cytoplasmic retention of MAFbx. Nucleocytoplasmic shuttling of MAFbx represents a novel mechanism for targeting its substrates and its cytosolic partners in muscle atrophy.

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Anti-diabetic and anti-obesity agent sodium tungstate enhances GCN pathway activation through Glc7p inhibition.

Publication Date: 2012 Feb 3 PMID: 22245679
Authors: Rodriguez-Hernandez, C. J. - Guinovart, J. J. - Murguia, J. R.
Journal: FEBS Lett

Tungstate counteracts diabetes and obesity in animal models, but its molecular mechanisms remain elusive. Our Saccharomyces cerevisiae-based approach has found that tungstate alleviated the growth defect induced by nutrient stress and enhanced the activation of the GCN pathway. Tungstate relieved the sensitivity to starvation of a gcn2-507 yeast hypomorphic mutant, indicating that tungstate modulated the GCN pathway downstream of Gcn2p. Interestingly, tungstate inhibited Glc7p and PP1 phosphatase activity, both negative regulators of the GCN pathway in yeast and humans, respectively. Accordingly, overexpression of a dominant-negative Glc7p mutant in yeast mimicked tungstate effects. Therefore tungstate alleviates nutrient stress in yeast by in vivo inhibition of Glc7p. These data uncover a potential role for tungstate in the treatment of PP1 and GCN related diseases.

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Exploring the mechanism of lipid transfer during biosynthesis of the acidic lipopeptide antibiotic CDA.

Publication Date: 2012 Feb 3 PMID: 22245678
Authors: Kraas, F. I. - Giessen, T. W. - Marahiel, M. A.
Journal: FEBS Lett

The non-ribosomally synthesized lipodepsipeptide CDA belongs to the group of acidic lipopeptide antibiotics, whose members feature a fatty acid side chain that strongly affects their antimicrobial activity. This study elucidates the N-acylation of the N-terminal serine in the CDA peptide chain. This reaction is referred to as lipoinitiation and is shown to be catalyzed by the dissected starter C domain found at the N-terminus of Cda-PSI. The recombinantly produced C domain specifically interacts with 2,3-epoxyhexanoyl-S-ACP and catalyzes the transfer of the fatty acid moiety onto the amino group of PCP-bound serine with high selectivity for both carrier protein bound substrates at the donor and acceptor site.

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Identification and characterization of an LCAT-like Arabidopsis thaliana gene encoding a novel phospholipase A.

Publication Date: 2012 Jan 10 PMID: 22245677
Authors: Chen, G. - Greer, M. S. - Lager, I. - Lindberg Yilmaz, J. - Mietkiewska, E. - Carlsson, A. S. - Stymne, S. - Weselake, R. J.
Journal: FEBS Lett

A previously uncharacterized Arabidopsis lecithin:cholesterol acyltransferase (LCAT) family gene (At4g19860) was functionally expressed in yeast, where it was demonstrated to encode a novel cytosolic and calcium-independent phospholipase A with preferences for the sn-2 position. This enzyme shows optimal activity at pH 5.0, exhibits a headgroup specificity for phosphatidylcholine>phosphatidic acid>phosphatidylethanolamine>phosphatidylglycerol>phosphatidylserine and has an acyl chain specificity for oleoyl>linoleoyl>ricinoleoyl. The expression of AtLCAT-PLA inhibited yeast cell growth and fatty acid accumulation. AtLCAT-PLA transcript in Arabidopsis was detected at high levels in roots and siliques.

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Inflammatory changes in adipose tissue enhance expression of GPR84, a medium-chain fatty acid receptor TNFalpha enhances GPR84 expression in adipocytes.

Publication Date: 2012 Jan 10 PMID: 22245676
Authors: Nagasaki, H. - Kondo, T. - Fuchigami, M. - Hashimoto, H. - Sugimura, Y. - Ozaki, N. - Arima, H. - Ota, A. - Oiso, Y. - Hamada, Y.
Journal: FEBS Lett

In this study we aimed to identify the physiological roles of G protein-coupled receptor 84 (GPR84) in adipose tissue, together with medium-chain fatty acids (MCFAs), the specific ligands for GPR84. In mice, high-fat diet up-regulated GPR84 expression in fat pads. In 3T3-L1 adipocytes, co-culture with a macrophage cell line, RAW264, or TNFalpha remarkably enhanced GPR84 expression. In the presence of TNFalpha, MCFAs down-regulated adiponectin mRNA expression in 3T3-L1 adipocytes. Taken together, our results suggest that GPR84 emerges in adipocytes in response to TNFalpha from infiltrating macrophages and exacerbates the vicious cycle between adiposity and diabesity.

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VapB as a regulator of osteoclastogenesis via modulation of PLCgamma2-Ca(2+)-NFAT signaling.

Publication Date: 2012 Feb 3 PMID: 22245675
Authors: Choi, S. W. - Yeon, J. T. - Park, K. I. - Lee, C. H. - Youn, B. S. - Oh, J. - Lee, M. S.
Journal: FEBS Lett

VapB has been shown to regulate calcium homeostasis in amyotrophic lateral sclerosis. Calcium signaling is also important in metabolic bone diseases, but the role of VapB in the generation of osteoclasts for bone resorption during osteoclastogenesis is not known. Therefore, we investigated the role of VapB in RANKL-induced osteoclast differentiation. Interestingly, VapB is induced during osteoclastogenesis, and regulates osteoclast differentiation by modulating NFATc1. The results also suggest that VapB regulates osteoclastogenesis via PLCgamma2-Ca(2+)-NFAT signaling. The involvement of PLCgamma2-Ca(2+)-NFAT signaling in VapB-regulated osteoclastogenesis was confirmed by a pharmacological study. Taken together, the results indicate that VapB positively regulates RANKL-mediated osteoclastogenesis via PLCgamma2-Ca(2+)-NFAT signaling.

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Sorbitol dehydrogenase of Aspergillus niger, SdhA, is part of the oxido-reductive d-galactose pathway and essential for d-sorbitol catabolism.

Publication Date: 2012 Jan 10 PMID: 22245674
Authors: Koivistoinen, O. M. - Richard, P. - Penttila, M. - Ruohonen, L. - Mojzita, D.
Journal: FEBS Lett

In filamentous fungi d-galactose can be catabolised through the oxido-reductive and/or the Leloir pathway. In the oxido-reductive pathway d-galactose is converted to d-fructose in a series of steps where the last step is the oxidation of d-sorbitol by an NAD-dependent dehydrogenase. We identified a sorbitol dehydrogenase gene, sdhA (JGI53356), in Aspergillus niger encoding a medium chain dehydrogenase which is involved in d-galactose and d-sorbitol catabolism. The gene is upregulated in the presence of d-galactose, galactitol and d-sorbitol. An sdhA deletion strain showed reduced growth on galactitol and growth on d-sorbitol was completely abolished. The purified enzyme converted d-sorbitol to d-fructose with K(m) of 50+/-5mM and v(max) of 80+/-10U/mg.

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Yeast ribosomal protein S3 possesses a beta-lyase activity on damaged DNA.

Publication Date: 2012 Jan 10 PMID: 22245673
Authors: Seong, K. M. - Jung, S. O. - Kim, H. D. - Kim, H. J. - Jung, Y. J. - Choi, S. Y. - Kim, J.
Journal: FEBS Lett

Yeast ribosomal protein S3 has multifunctional activities that are involved in both protein translation and DNA repair. Here, we report that yeast Rps3p cleaves variously damaged DNA that contains not only AP sites and pyrimidine dimers but also 7,8-hydro-8-oxoguanine. This study also revealed that Rps3p has a beta-lyase activity with a broad range of substrate specificity which cleaves phosphodiester bonds of UV or oxidatively damaged DNA substrates. Mutation analysis of the yeast Rps3 protein including introduction of domain deletions and residue replacements identified the residues Asp154 and Lys200 are important for the catalytic activity. In addition, the repair enzyme activity of yeast Rps3p was confirmed by complementation in xth, nfo Escherichia coli cells in which the DNA repair process is defective.

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PEDV ORF3 encodes an ion channel protein and regulates virus production.

Publication Date: 2012 Jan 11 PMID: 22245155
Authors: Wang, K. - Lu, W. - Chen, J. - Xie, S. - Shi, H. - Hsu, H. - Yu, W. - Xu, K. - Bian, C. - Fischer, W. B. - Schwarz, W. - Feng, L. - Sun, B.
Journal: FEBS Lett

Several studies suggest that the open reading frame 3 (ORF3) gene of porcine epidemic diarrhea virus (PEDV) is related to viral infectivity and pathogenicity, but its function remains unknown. Here, we propose a structure model of the ORF3 protein consisting of four TM domains and forming a tetrameric assembly. ORF3 protein can be detected in PEDV-infected cells and it functions as an ion channel in both Xenopus laevis oocytes and yeast. Mutation analysis showed that Tyr170 in TM4 is important for potassium channel activity. Furthermore, viral production is reduced in infected Vero cells when ORF3 gene is silenced by siRNA. Interestingly, the ORF3 gene from an attenuated PEDV encodes a truncated protein with 49 nucleotide deletions, which lacks the ion channel activity.

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Activation of macrophage-stimulating protein by human airway trypsin-like protease.

Publication Date: 2012 Feb 3 PMID: 22245154
Authors: Orikawa, H. - Kawaguchi, M. - Baba, T. - Yorita, K. - Sakoda, S. - Kataoka, H.
Journal: FEBS Lett

Macrophage-stimulating protein (MSP) circulates as a proform protein and requires proteolytic processing for activation. Respiratory ciliated cells express the MSP receptor, recepteur d'origine nantais (RON), at the apical surface, which reportedly has an important role in ciliary function. Like RON, human airway trypsin-like protease (HAT) is also expressed at the apical surface of ciliated cells. Here we show that HAT cleaves proMSP at the physiological activation site, Arg483-Val484. MSP processed by HAT could induce chemotactic responses and morphological changes of peritoneal macrophages. In human respiratory epithelial cells, knock down of HAT expression reduced proMSP processing and RON autophosphorylation. We suggest that HAT is important for MSP-RON signaling in the respiratory tract. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: HATcleavesproMSP by protease assay (View interaction).

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SNPs of hemocyanin C-terminal fragment in shrimp Litopenaeus vannamei.

Publication Date: 2012 Jan 11 PMID: 22245153
Authors: Zhao, X. - Guo, L. - Zhang, Y. - Liu, Y. - Zhang, X. - Lun, J. - Chen, J. - Li, Y.
Journal: FEBS Lett

In this study, we identified a variable region in the C-terminus of hemocyanin from the shrimp Litopenaeus vannamei (2288-2503bp, HcSC) by sequence alignments. A total of 13 SNPs were identified by PCR-SSCP and HcSC clone sequencing. The SSCP patterns of HcSC could be modulated in Vibro parahaemolyticus-treated shrimps. A novel SSCP band with four SNP sites was identified in V. parahaemolyticus-resistant shrimps. More importantly, three of these four SNPs introduced variations in amino acid sequence and possibly secondary structure of the HcSC polypeptide and resulted in a higher agglutinative activity against seven pathogenic bacteria. These results suggest that the C-terminus of shrimp L. vannamei hemocyanin possesses SNPs, which may be related to shrimp resistance to different pathogens.

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Hypoxia-induced SM22alpha in A549 cells activates the IGF1R/PI3K/Akt pathway, conferring cellular resistance against chemo- and radiation therapy.

Publication Date: 2012 Jan 10 PMID: 22245152
Authors: Kim, T. R. - Cho, E. W. - Paik, S. G. - Kim, I. G.
Journal: FEBS Lett

Chemo- or radiation-resistance in tumors caused by hypoxia often undermines efficacy of cancer therapy. Thus, therapies that overcome cellular resistance during hypoxia are necessary. SM22alpha is an actin-binding protein found in smooth muscle, fibroblasts, and some epithelium. We demonstrate that SM22alpha is induced in A549 non-small cell lung carcinoma cells by hypoxia and its overexpression increased chemo- and radiation-resistance. Hypoxia-mediated induction of SM22alpha expression is hypoxia-inducible factor-independent. Moreover, SM22alpha overexpression enhances tumor cell growth and activates the IGF1R/PI3K/Akt pathway via direct interaction with IGF1Rbeta. Our results suggest SM22alpha as a novel regulator of hypoxic survival pathway of A549 NSCLC cells. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: IGFR1 Betaphysically interacts with SM22 alpha by anti bait coimmunoprecipitation (View Interaction: 1, 2).

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Mutation of a raft-targeting signal in the transmembrane region retards transport of influenza virus hemagglutinin through the Golgi.

Publication Date: 2012 Feb 3 PMID: 22245151
Authors: Engel, S. - de Vries, M. - Herrmann, A. - Veit, M.
Journal: FEBS Lett

Inclusion of proteins into membrane-rafts favours interactions required for virus assembly but has also been proposed to facilitate vesicular transport of proteins. The hemagglutinin (HA) of influenza virus contains a raft-targeting sequence in the outer leaflet of its transmembrane region. We report that its mutation enhances co-localization of HA with a cis-Golgi marker and retards Golgi-localized processing, such as acquisition of Endo-H resistant carbohydrates and proteolytic cleavage. In contrast, trimerization of the molecule in the ER and transport to the apical membrane were not affected. The second signal for raft-targeting, S-acylation at cytoplasmic cysteines, did not retard HA transport.

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Recognition of different DNA sequences by a DNA-binding protein alters protein dynamics differentially.

Publication Date: 2012 Feb 3 PMID: 22240202
Authors: Mondol, T. - Batabyal, S. - Mazumder, A. - Roy, S. - Pal, S. K.
Journal: FEBS Lett

lambda-Repressor-operator sites interaction, particularly O(R)1 and O(R)2, is a key component of the lambda-genetic switch. FRET from the dansyl bound to the C-terminal domain of the protein, to the intercalated EtBr in the operator DNA indicates that the structure of the protein is more compact in the O(R)2 complex than in the O(R)1 complex. Fluorescence anisotropy reveals enhanced flexibility of the C-terminal domain of the repressor at fast timescales after complex formation with O(R)1. In contrast, O(R)2 bound repressor shows no significant enhancement of protein dynamics at these timescales. These differences are shown to be important for correct protein-protein interactions. Altered protein dynamics upon specific DNA sequence recognition may play important roles in assembly of regulatory proteins at the correct positions.

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Human lactoferrin suppresses TNF-alpha-induced intercellular adhesion molecule-1 expression via competition with NF-kappaB in endothelial cells.

Publication Date: 2012 Feb 3 PMID: 22226679
Authors: Kim, C. W. - Lee, T. H. - Park, K. H. - Choi, S. Y. - Kim, J.
Journal: FEBS Lett

Lactoferrin (Lf) is known to have anti-inflammatory activity, but the mechanisms of action by Lf remain to be elucidated. Here, we demonstrated that TNF-alpha-induced expression of intercellular adhesion molecule-1 (ICAM-1) was down-regulated by Lf in a DNA-binding dependent manner at transcriptional level in endothelial cells. Our results showed that Lf bound to a DNA region in the ICAM-1 promoter in vitro as well as in chromatin context. Lf inhibited binding of NF-kappaB to a proximal NF-kappaB site in ICAM-1 promoter. This type of repression represents an additional mechanism for the action of Lf in regulation of gene expression.

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C-terminal residues of Oryza sativa GUN4 are required for the activation of the ChlH subunit of magnesium chelatase in chlorophyll synthesis.

Publication Date: 2012 Feb 3 PMID: 22226678
Authors: Zhou, S. - Sawicki, A. - Willows, R. D. - Luo, M.
Journal: FEBS Lett

Oryza sativa GUN4 together with the magnesium chelatase subunits ChlI, ChlD, and ChlH have been heterologously expressed and purified to reconstitute magnesium chelatase activity in vitro. Maximum magnesium chelatase activity requires pre-activation of OsChlH with OsGUN4, Mg(2+) and protoporphyrin-IX. OsGUN4 and OsChlH preincubated without protoporphyrin-IX yields magnesium chelatase activity similar to assays without OsGUN4, suggesting formation of a dead-end complex. Either 9 or 10 C-terminal amino acids of OsGUN4 are slowly hydrolyzed to yield a truncated OsGUN4. These truncated OsGUN4 still bind protoporphyrin-IX and Mg-protoporphyrin-IX but are unable to activate OsChlH. This suggests the mechanism of GUN4 activation of magnesium chelatase is different in eukaryotes compared to cyanobacteria as the orthologous cyanobacterial GUN4 proteins lack this C-terminal extension. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: ChlH and ChlHbind by molecular sieving (View interaction) ChlH and ChlHbind by molecular sieving (View interaction) ChlD and ChlDbind by molecular sieving (View interaction).

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Structural and biochemical insight into glycogenin inactivation by the glycogenosis-causing T82M mutation.

Publication Date: 2012 Feb 3 PMID: 22226635
Authors: Carrizo, M. E. - Romero, J. M. - Issoglio, F. M. - Curtino, J. A.
Journal: FEBS Lett

The X-ray structure of rabbit glycogenin containing the T82M (T83M according to previous authors amino acid numbering [1]) mutation causing glycogenosis showed the loss of Thr82 hydrogen bond to Asp162, the residue involved in the activation step of the glucose transfer reaction mechanism. Autoglucosylation, maltoside transglucosylation and UDP-glucose hydrolyzing activities were abolished even though affinity and interactions with UDP-glucose and positioning of Tyr194 acceptor were conserved. Substitution of Thr82 for serine but not for valine restored the maximum extent of autoglucosylation as well as transglucosylation and UDP-glucose hydrolysis rate. Results provided evidence sustaining the essential role of the lost single hydrogen bond for UDP-glucose activation leading to glycogenin-bound glycogen primer synthesis.

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FLU, a negative feedback regulator of tetrapyrrole biosynthesis, is physically linked to the final steps of the Mg(++)-branch of this pathway.

Publication Date: 2012 Feb 3 PMID: 22212719
Authors: Kauss, D. - Bischof, S. - Steiner, S. - Apel, K. - Meskauskiene, R.
Journal: FEBS Lett

Regulation of tetrapyrrole biosynthesis in higher plants has been attributed to negative feedback control. Two effectors of feedback inhibition have been identified, heme and the FLU protein. Inhibition by heme implicates the Fe-branch via regulation of the initial step of tetrapyrrole synthesis. In the present work a FLU-containing chloroplast membrane complex was identified, that besides FLU comprises the four enzymes catalyzing the final steps of chlorophyll synthesis. The results support the notion that FLU links chlorophyll synthesis and the target of feedback control, glutamyl-tRNA reductase, thereby allowing also the Mg-branch to control the initial step of tetrapyrrole synthesis. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: FLU, CHL27, PORB, PORC and CHLPphysically interact by blue native page (View interaction) FLUphysically interacts with CHL27, PORB, PORC, CHLP and GluTR by anti tag co-immunoprecipitation (View interaction) FLUphysically interacts with CHL27 by anti bait co-immunoprecipitation (View Interaction: 1, 2) FLUphysically interacts with CHL27 by anti tag co-immunoprecipitation (View interaction) FLUphysically interacts with CHL27, PORB, PORC and CHLP by anti tag co-immunoprecipitation (View interaction).

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Functional characterisation of human SGLT-5 as a novel kidney-specific sodium-dependent sugar transporter.

Publication Date: 2012 Feb 3 PMID: 22212718
Authors: Grempler, R. - Augustin, R. - Froehner, S. - Hildebrandt, T. - Simon, E. - Mark, M. - Eickelmann, P.
Journal: FEBS Lett

Sodium glucose cotransporters (SGLT) actively catalyse carbohydrate transport across cellular membranes. Six of the 12 known SGLT family members have the capacity to bind and/or transport monosaccharides (SGLT-1 to 6); of these, all but SGLT-5 have been characterised. Here we demonstrate that human SGLT-5 is exclusively expressed in the kidney. Four splice variants were detected and the most abundant SGLT-5-mRNA was functionally characterised. SGLT-5 mediates sodium-dependent [(14)C]-alpha-methyl-d-glucose (AMG) transport that can be inhibited by mannose, fructose, glucose, and galactose. Uptake studies using demonstrated high capacity transport for mannose and fructose and, to a lesser extent, glucose, AMG, and galactose. SGLT-5 mediated mannose, fructose and AMG transport was weakly (muM potency) inhibited by SGLT-2 inhibitors. In summary, we have characterised SGLT-5 as a kidney mannose transporter. Further studies are warranted to explore the physiological role of SGLT-5.

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Phosphorylation of Akt/GSK-3beta/eNOS amplifies 5-HT(2B) receptor blockade mediated anti-hypertrophic effect in rats.

Publication Date: 2012 Jan 20 PMID: 22210189
Authors: Bharti, S. - Singh, R. - Chauhan, S. S. - Hussain, T. - Al-Attas, O. S. - Arya, D. S.
Journal: FEBS Lett

Herein, we studied the cross talk between 5-HT(2B) receptor blocker (SB-204741) and GSK-3beta inhibitor (SB-216763) in isoproterenol-induced cardiac hypertrophy for 28days. SB-204741 treatment significantly ameliorated (P<0.05) myocardial dysfunction, myocyte area, fibrosis and myocardial architecture in isoproterenol insulted myocardium. Moreover, this improvement in functional and morphological changes was associated with suppression of hypertrophic (BNP and CK-MB), inflammatory (IKK-beta/NF-kappaB/TNF-alpha and CRP), and apoptotic markers (TUNEL positivity and Bax expression) along with phosphorylation of Akt/GSK-3beta/beta-catenin/eNOS. Intriguingly, co-treatment with GSK-3beta inhibitor (P<0.01) further amplified the anti-hypertrophic effect of SB-204741 (P<0.05) such that the effect was indistinguishable from that of vehicle treated rats. Thus, 5-HT(2B) receptor blockade mediated anti-hypertrophic effect is atleast in part is governed through phosphorylation of Akt/GSK-3beta/beta-catenin/eNOS via attenuating inflammatory and apoptotic pathways.

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Interaction of protein inhibitor of activated STAT 2 (PIAS2) with receptor of activated C kinase 1, RACK1.

Publication Date: 2012 Jan 20 PMID: 22210188
Authors: Zheng, Y. - Zhang, L. - Jia, X. - Wang, H. - Hu, Y.
Journal: FEBS Lett

In this study, the evolutionarily conserved intracellular adaptor protein, receptor of activated C kinase 1 (RACK1) was identified as a novel interaction partner of protein inhibitor of activated STAT 2 (PIAS2) using a yeast two-hybrid screening system. The direct interaction and co-localization of RACK1 with PIAS2 was confirmed by immunoprecipitation and immunofluorescence staining analysis, respectively. The 5th to 7th Trp-Asp 40 (5-7 WD40) repeats of RACK1 were identified as the minimal domain required for interaction with PIAS2 by deletion analysis. Furthermore, multiple PIAS2-domains, particularly the 'PINIT' and RLD domains, bind the RACK1 5-7 WD40 domain. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: PIAS2physically interacts with RACK1 by two hybrid (View interaction) RACK1 and PIAS2colocalize by fluorescence microscopy (View interaction) PIAS2physically interacts with RACK1 by two hybrid pooling approach (View interaction) PIAS2 and RACK1colocalize by fluorescence microscopy (View interaction) PIAS2physically interacts with RACK1 by anti bait coimmunoprecipitation (View Interaction: 1, 2).

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Mitochondria targeting of non-peroxidizable triphenylphosphonium conjugated oleic acid protects mouse embryonic cells against apoptosis: Role of cardiolipin remodeling.

Publication Date: 2012 Feb 3 PMID: 22210054
Authors: Tyurina, Y. Y. - Tungekar, M. A. - Jung, M. Y. - Tyurin, V. A. - Greenberger, J. S. - Stoyanovsky, D. A. - Kagan, V. E.
Journal: FEBS Lett

Peroxidation of cardiolipin in mitochondria is essential for the execution of apoptosis. We suggested that integration of oleic acid into cardiolipin generates non-oxidizable cardiolipin species hence protects cells against apoptosis. We synthesized mitochondria-targeted triphenylphosphonium oleic acid ester. Using lipidomics analysis we found that pretreatment of mouse embryonic cells with triphenylphosphonium oleic acid ester resulted in decreased contents of polyunsaturated cardiolipins and elevation of its species containing oleic acid residues. This caused suppression of apoptosis induced by actinomycin D. Triacsin C, an inhibitor of acyl-CoA synthase, blocked integration of oleic acid into cardiolipin and restored cell sensitivity to apoptosis.

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Double-stranded RNA induces S100 gene expression by a cycloheximide-sensitive factor.

Publication Date: 2012 Jan 20 PMID: 22209981
Authors: Voss, A. - Gescher, K. - Hensel, A. - Nacken, W. - Zanker, K. S. - Kerkhoff, C.
Journal: FEBS Lett

Viral double-stranded RNA (dsRNA) and its synthetic analog polyI:C are recognized via multiple pathways and induce the expression of genes related to inflammation. In the present study, we demonstrated the polyI:C-induced gene expression of the damage associated molecular pattern (DAMP) molecules S100A8 and S100A9, while other S100 genes were not affected. Cycloheximide and Brefeldin A treatment revealed both the expression of S100A8 and S100A9 as secondary response genes and the involvement of polyI:C-induced cytokines herein. Several type I and type III interferons such as IFNbeta, IL-20, IL-24, and IFNlambda/IL-29 were expressed in response to polyI:C, however, they failed to induce S100A8 and S100A9 gene expression. These data indicate the involvement of the danger molecule S100A8/A9 in the resistance against viruses.

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Splice blocking of zygotic sox31 leads to developmental arrest shortly after Mid-Blastula Transition and induces apoptosis in zebrafish.

Publication Date: 2012 Feb 3 PMID: 22209980
Authors: Hu, S. N. - Yu, H. - Zhang, Y. B. - Wu, Z. L. - Yan, Y. C. - Li, Y. X. - Li, Y. Y. - Li, Y. P.
Journal: FEBS Lett

Here we report that splice blocking morpholinos (Sb MO) against zebrafish sox31 elicit developmental arrest, likely through creating a series of dominant negative splicing variants. Embryos injected with the Sb MO develop normally before the Mid-Blastula Transition (MBT); however, they do not initiate epiboly. Microarray analysis of mRNAs collected at the dome stage revealed that the Sb MO impairs activation of a large set of zygotic genes and reduces degradation of maternal mRNA during MBT. Furthermore, an apoptotic response occurs in Sb morphants at about 6hpf. SoxB1 family genes including sox31 thus play an essential role for early embryos traversing the transitional stage.

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Chinese hamster AP endonuclease operates by a two-metal ion assisted catalytic mechanism.

Publication Date: 2012 Feb 3 PMID: 22209979
Authors: Borjigin, M. - Arenaz, P. - Stec, B.
Journal: FEBS Lett

The APE1, an important mammalian AP endonuclease, is an essential enzyme in the base excision DNA repair pathway (BER). The number of metal ions involved directly in the catalysis remains controversial. Here we describe the metal ion titration experiments that demonstrate the requirement for two metal ions for the endonuclease activity of the Chinese hamster APE1. The titration with the non-activating metal ion La(3+) showed a biphasic behavior with activating and inhibitory effects of La(3+) in the range of 0-100muM in the presence of 5mMMg(2+). Modeling of the enzyme-substrate/product complexes provided insight into the endonuclease activity and elucidated the nature of the crystal structures. Accordingly, we proposed a reaction scheme for the two-metal ion assisted catalysis of chAPE1 endonuclease activity.

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Formation of supramolecular structures of a native-like protein in the presence of amphiphilic peptides: Variations in aggregate morphology.

Publication Date: 2012 Jan 20 PMID: 22200573
Authors: Artemova, N. - Stein-Margolina, V. - Smirnova, E. - Gurvits, B.
Journal: FEBS Lett

A striking potential of the amphiphilic dipeptides, Arg-Phe or Asp-Phe, to induce aggregation of a model protein, alcohol dehydrogenase in its native-like state, has been demonstrated under physiologically relevant conditions, using dynamic light scattering, fluorescence spectroscopy, circular dichroism, transmission electron- and atomic force microscopy. The peptide action resulted in accumulation of a variety of morphologically distinct supramolecular structures profoundly differing from those generated by the heat-induced aggregation at the early stages of the process, when amyloid fibril assemblies were not detectable. The biogenic amphiphilic agents are suggested to act as regulators of structural transformations of native-like proteins.

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A novel family of toxin/antitoxin proteins in Bacillus species.

Publication Date: 2012 Jan 20 PMID: 22200572
Authors: Holberger, L. E. - Garza-Sanchez, F. - Lamoureux, J. - Low, D. A. - Hayes, C. S.
Journal: FEBS Lett

The C-terminal regions (CT) of Pfam PF04740 proteins share significant sequence identity with the toxic CdiA-CT effector domains of contact-dependent growth inhibition (CDI) systems. In accord with this homology, we find that several PF04740 CT domains inhibit cell growth when expressed in Escherichia coli. This growth inhibition is specifically blocked by antitoxin proteins encoded downstream of each PF04740 gene. The YobL-CT, YxiD-CT and YqcG-CT domains from Bacillus subtilis 168 have cytotoxic RNase activities, which are neutralized by the binding of cognate YobK, YxxD and YqcF antitoxin proteins, respectively. Our results show that PF04740 proteins represent a new family of toxin/antitoxin pairs that are widely distributed in Gram-positive bacteria. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: YobKbinds to YobL by pull down (View interaction) YezGbinds to YeeF by pull down (View interaction).

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Dissociation of the insulin receptor from caveolae during TNFalpha-induced insulin resistance and its recovery by d-PDMP.

Publication Date: 2012 Jan 20 PMID: 22200571
Authors: Sekimoto, J. - Kabayama, K. - Gohara, K. - Inokuchi, J.
Journal: FEBS Lett

Previously, we demonstrated that an inhibitor of ganglioside biosynthesis, d-PDMP, could restore impaired insulin signaling in tumor necrosis factor alpha (TNFalpha)-treated adipocytes by blocking the increase of GM3 ganglioside. Here, we analyzed the interaction between insulin receptor (IR) and GM3 in the plasma membranes using immunoelectron microscopy. In normal adipocytes, most GM3 molecules localized at planar and non-caveolar regions. Approximately 19% of IR molecules were detected in caveolar regions. The relative ratio of IRs associated with caveolae in TNFalpha-treated adipocytes was decreased to one-fifth of that in normal adipocytes, but this decrease was restored by d-PDMP. Thus, we could obtain direct evidence that insulin resistance is a membrane microdomain disorder caused by aberrant expression of ganglioside.

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Effect of N-homocysteinylation on physicochemical and cytotoxic properties of amyloid beta-peptide.

Publication Date: 2012 Jan 20 PMID: 22200570
Authors: Khodadadi, S. - Riazi, G. H. - Ahmadian, S. - Hoveizi, E. - Karima, O. - Aryapour, H.
Journal: FEBS Lett

Abstract Hyperhomocysteinemia has recently been identified as an important risk factor for Alzheimer's disease (AD). One of the potential mechanisms underlying harmful effects of homocysteine (Hcy) is site-specific acylation of proteins at lysine residues by homocysteine thiolactone (HCTL). The accumulation of amyloid beta-peptide (Abeta) in the brain is a neuropathological hallmark of AD. In the present study we were interested to investigate the effects of N-homocysteinylation on the aggregation propensity and neurotoxicity of Abeta(1-42). By coupling several techniques, we demonstrated that the homocysteinylation of lysine residues increase the neurotoxicity of the Abeta peptide by stabilizing soluble oligomeric intermediates. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: A Beta 1-42 and A Beta 1-42bind by fluorescence technology (View interaction) A Beta 1-42 and A Beta 1-42bind by electron microscopy (View interaction).

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Comparing the temperature dependence of FMN to heme electron transfer in full length and truncated inducible nitric oxide synthase proteins.

Publication Date: 2012 Jan 20 PMID: 22198200
Authors: Li, W. - Chen, L. - Fan, W. - Feng, C.
Journal: FEBS Lett

The FMN-heme interdomain (intraprotein) electron transfer (IET) kinetics in full length and oxygenase/FMN (oxyFMN) construct of human iNOS were determined by laser flash photolysis over the temperature range from 283 to 304K. An appreciable increase in the rate constant value was observed with an increase in the temperature. Our previous viscosity study indicated that the IET process is conformationally gated, and Eyring equation was thus used to analyze the temperature dependence data. The obtained magnitude of activation entropy for the IET in the oxyFMN construct is only one-fifth of that for the holoenzyme. This indicates that the FMN domain in the holoenzyme needs to sample more conformations before the IET takes place, and that the FMN domain in the oxyFMN construct is better poised for efficient IET.

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The cytosolic domain of human Tom22 modulates human Bax mitochondrial translocation and conformation in yeast.

Publication Date: 2012 Jan 20 PMID: 22198199
Authors: Renault, T. T. - Grandier-Vazeille, X. - Arokium, H. - Velours, G. - Camougrand, N. - Priault, M. - Teijido, O. - Dejean, L. M. - Manon, S.
Journal: FEBS Lett

The role of the mitochondrial protein receptor Tom22p in the interaction of pro-apoptotic protein Bax with yeast mitochondria was investigated. Co-immunoprecipitation assays showed that human Bax interacted with different TOM subunits, including Tom22p. Expression of the cytosolic receptor domain of human Tom22 increased Bax mitochondrial localization, but decreased the proportion of active Bax. BN-PAGE showed that the cytosolic domain of Tom22 interfered with the oligomerization of Bax. These data suggest that the interaction with the cytosolic domain of Tom22 helps Bax to acquire a conformation able to interact with the outer mitochondrial membrane. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: BAX and BAXphysically interact with TOM22 by anti bait coimmunoprecipitation (View interaction) BAXphysically interacts with TOM70 by anti bait coimmunoprecipitation (View interaction) BAXphysically interacts with TOM22, TOM70 and TOM40 by anti bait coimmunoprecipitation (View interaction) BAXphysically interacts with TOM22 by anti bait coimmunoprecipitation (View interaction) BAX and BAXbind by blue native page (View interaction) BAXphysically interacts with TOM40 by anti bait coimmunoprecipitation (View interaction).

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Lifetimes of photosystem I and II proteins in the cyanobacterium Synechocystis sp. PCC 6803.

Publication Date: 2012 Jan 20 PMID: 22197103
Authors: Yao, D. C. - Brune, D. C. - Vermaas, W. F.
Journal: FEBS Lett

The half-life times of photosystem I and II proteins were determined using (15)N-labeling and mass spectrometry. The half-life times (30-75h for photosystem I components and <1-11h for the large photosystem II proteins) were similar when proteins were isolated from monomeric vs. oligomeric complexes on Blue-Native gels, suggesting that the two forms of both photosystems can interchange on a timescale of <1h or that only one form of each photosystem exists in thylakoids in vivo. The half-life times of proteins associated with either photosystem generally were unaffected by the absence of Small Cab-like proteins.

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Geranyl modification on the tryptophan residue of ComX(RO-E-2) pheromone by a cell-free system.

Publication Date: 2012 Jan 20 PMID: 22197102
Authors: Tsuji, F. - Ishihara, A. - Kurata, K. - Nakagawa, A. - Okada, M. - Kitamura, S. - Kanamaru, K. - Masuda, Y. - Murakami, K. - Irie, K. - Sakagami, Y.
Journal: FEBS Lett

ComX pheromone is an isoprenoidal oligopeptide containing a modified tryptophan residue, which stimulates natural genetic competence in the gram-positive bacterium Bacillus. Since posttranslational prenylation on the tryptophan residue has not been reported except in ComX pheromone, the universality of this modification has not yet been elucidated. In this paper, we established a cell-free system, whereby the tryptophan residue in peptides is modified with a geranyl group by modifying enzyme ComQ. In addition, we investigated enzymatic reaction conditions using an in vitro enzyme reaction system. This is the first report of in vitro geranylation on the tryptophan residue. This system is potentially a useful tool for elucidating the universality of prenylation on the tryptophan residue.

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14-3-3sigma regulation by p53 mediates a chemotherapy response to 5-fluorouracil in MCF-7 breast cancer cells via Akt inactivation.

Publication Date: 2012 Jan 20 PMID: 22192357
Authors: Zheng, G. - Xiong, Y. - Yi, S. - Zhang, W. - Peng, B. - Zhang, Q. - He, Z.
Journal: FEBS Lett

We previously demonstrated that 14-3-3sigma was downregulated in 5-fluorouracil (5-Fu)-resistant MCF-7 breast cancer cells (MCF-7/5-Fu). Here, we found that stably enhanced 14-3-3sigma expression strengthened the effects of 5-Fu, Mitoxantrone and cDDP. 14-3-3sigma stabilised the p53 protein and bound Akt to inhibit its activity and its downstream targets: survivin, Bcl-2 and NF-kappaB-p50. In addition, decreased p53 expression, but not promoter hypermethylation, was responsible for the downregulation of 14-3-3sigma in MCF-7/5-Fu cells. Meanwhile, initial treatments with high concentrations of 5-Fu clearly induced 14-3-3sigma and p53 expression in a time-dependent manner. 14-3-3sigma-mediated molecular events that synergise with p53 may play important roles in the chemotherapy of breast cancer.

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Specific nitration of tyrosines 46 and 48 makes cytochrome c assemble a non-functional apoptosome.

Publication Date: 2012 Jan 20 PMID: 22192356
Authors: Garcia-Heredia, J. M. - Diaz-Moreno, I. - Diaz-Quintana, A. - Orzaez, M. - Navarro, J. A. - Hervas, M. - De la Rosa, M. A.
Journal: FEBS Lett

Under nitroxidative stress, a minor fraction of cytochrome c can be modified by tyrosine nitration. Here we analyze the specific effect of nitration of tyrosines 46 and 48 on the dual role of cytochrome c in cell survival and cell death. Our findings reveal that nitration of these two solvent-exposed residues has a negligible effect on the rate of electron transfer from cytochrome c to cytochrome c oxidase, but impairs the ability of the heme protein to activate caspase-9 by assembling a non-functional apoptosome. It seems that cytochrome c nitration under cellular stress counteracts apoptosis in light of the small amount of modified protein. We conclude that other changes such as increased peroxidase activity prevail and allow the execution of apoptosis.

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cGMP and cyclic nucleotide-gated channels participate in mouse sperm capacitation.

Publication Date: 2012 Jan 20 PMID: 22192355
Authors: Cisneros-Mejorado, A. - Sanchez Herrera, D. P.
Journal: FEBS Lett

During capacitation of mammalian sperm intracellular [Ca(2+)] and cyclic nucleotides increase, suggesting that CNG channels play a role in the physiology of sperm. Here we study the effect of capacitation, 8Br-cAMP (8-bromoadenosine 3',5'-cyclic monophosphate) and 8Br-cGMP (8-bromoguanosine 3',5'-cyclic monophosphate) on the macroscopic ionic currents of mouse sperm, finding the existence of different populations of sperm, in terms of the recorded current and its response to cyclic nucleotides. Our results show that capacitation and cyclic nucleotides increase the ionic current, having a differential sensitivity to cGMP (cyclic guanosine monophosphate) and cAMP (cyclic adenosine monophosphate). Using a specific inhibitor we determine the contribution of CNG channels to macroscopic current and capacitation.

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Discovery of a novel enzymatic cleavage site for botulinum neurotoxin F5.

Publication Date: 2012 Jan 20 PMID: 22172278
Authors: Kalb, S. R. - Baudys, J. - Webb, R. P. - Wright, P. - Smith, T. J. - Smith, L. A. - Fernandez, R. - Raphael, B. H. - Maslanka, S. E. - Pirkle, J. L. - Barr, J. R.
Journal: FEBS Lett

Botulinum neurotoxins (BoNTs) cause botulism by cleaving proteins necessary for nerve transmission. There are seven serotypes of BoNT, A-G, characterized by their response to antisera. Many serotypes are further distinguished into differing subtypes based on amino acid sequence, some of which result in functional differences. Our laboratory previously reported that all tested subtypes within each serotype have the same site of enzymatic activity. Recently, three new subtypes of BoNT/F; /F3, /F4, and /F5, were reported. Here, we report that BoNT/F5 cleaves substrate synaptobrevin-2 in a different location than the other BoNT/F subtypes, between (54)L and (55)E. This is the first report of cleavage of synaptobrevin-2 in this location. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: BoNT/F5cleavesSynaptobrevin-2 by protease assay (View interaction: 1, 2) BoNT/F1cleavesSynaptobrevin-2 by protease assay (View interaction: 1, 2).

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The yeast metacaspase is implicated in oxidative stress response in frataxin-deficient cells.

Publication Date: 2012 Jan 20 PMID: 22155640
Authors: Lefevre, S. - Sliwa, D. - Auchere, F. - Brossas, C. - Ruckenstuhl, C. - Boggetto, N. - Lesuisse, E. - Madeo, F. - Camadro, J. M. - Santos, R.
Journal: FEBS Lett

Friedreich ataxia is the most common recessive neurodegenerative disease and is caused by reduced expression of mitochondrial frataxin. Frataxin depletion causes impairment in iron-sulfur cluster and heme biosynthesis, disruption of iron homeostasis and hypersensitivity to oxidants. Currently no pharmacological treatment blocks disease progression, although antioxidant therapies proved to benefit patients. We show that sensitivity of yeast frataxin-deficient cells to hydrogen peroxide is partially mediated by the metacaspase. Metacaspase deletion in frataxin-deficient cells results in recovery of antioxidant capacity and heme synthesis. In addition, our results suggest that metacaspase is associated with mitochondrial respiration, intracellular redox control and genomic stability.

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Acetylation-dependent regulation of mitochondrial ALDH2 activation by SIRT3 mediates acute ethanol-induced eNOS activation.

Publication Date: 2012 Jan 20 PMID: 22155639
Authors: Xue, L. - Xu, F. - Meng, L. - Wei, S. - Wang, J. - Hao, P. - Bian, Y. - Zhang, Y. - Chen, Y.
Journal: FEBS Lett

Moderate alcohol consumption has beneficial effects on endothelial nitric-oxide synthase (eNOS) activation, which can engender an array of anti-atherogenic actions. Here we show that in human aortic endothelial cells (HAECs), rapid activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) mediates ethanol-induced eNOS activation by preventing reactive oxygen species (ROS) accumulation. Furthermore, activation of ALDH2 by ethanol is due to its hyperacetylation by SIRT3 inactivation. These data suggest that ethanol-induced eNOS activation in HAECs may be dependent on ALDH2 hyperacetylation by SIRT3 inactivation.

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