Developmental Cell

Differential H3K4 methylation identifies developmentally poised hematopoietic genes.

Publication Date: 2008 May PMID: 18477461
Authors: Orford, K. - Kharchenko, P. - Lai, W. - Dao, M. C. - Worhunsky, D. J. - Ferro, A. - Janzen, V. - Park, P. J. - Scadden, D. T.
Journal: Dev Cell

Throughout development, cell fate decisions are converted into epigenetic information that determines cellular identity. Covalent histone modifications are heritable epigenetic marks and are hypothesized to play a central role in this process. In this report, we assess the concordance of histone H3 lysine 4 dimethylation (H3K4me2) and trimethylation (H3K4me3) on a genome-wide scale in erythroid development by analyzing pluripotent, multipotent, and unipotent cell types. Although H3K4me2 and H3K4me3 are concordant at most genes, multipotential hematopoietic cells have a subset of genes that are differentially methylated (H3K4me2+/me3-). These genes are transcriptionally silent, highly enriched in lineage-specific hematopoietic genes, and uniquely susceptible to differentiation-induced H3K4 demethylation. Self-renewing embryonic stem cells, which restrict H3K4 methylation to genes that contain CpG islands (CGIs), lack H3K4me2+/me3- genes. These data reveal distinct epigenetic regulation of CGI and non-CGI genes during development and indicate an interactive relationship between DNA sequence and differential H3K4 methylation in lineage-specific differentiation.

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Myosin phosphatase-targeting subunit 1 regulates mitosis by antagonizing polo-like kinase 1.

Publication Date: 2008 May PMID: 18477460
Authors: Yamashiro, S. - Yamakita, Y. - Totsukawa, G. - Goto, H. - Kaibuchi, K. - Ito, M. - Hartshorne, D. J. - Matsumura, F.
Journal: Dev Cell

Myosin phosphatase-targeting subunit 1 (MYPT1) binds to the catalytic subunit of protein phosphatase 1 (PP1C). This binding is believed to target PP1C to specific substrates including myosin II, thus controlling cellular contractility. Surprisingly, we found that during mitosis, mammalian MYPT1 binds to polo-like kinase 1 (PLK1). MYPT1 is phosphorylated during mitosis by proline-directed kinases including cdc2, which generates the binding motif for the polo box domain of PLK1. Depletion of PLK1 by small interfering RNAs is known to result in loss of gamma-tubulin recruitment to the centrosomes, blocking centrosome maturation and leading to mitotic arrest. We found that codepletion of MYPT1 and PLK1 reinstates gamma-tubulin at the centrosomes, rescuing the mitotic arrest. MYPT1 depletion increases phosphorylation of PLK1 at its activating site (Thr210) in vivo, explaining, at least in part, the rescue phenotype by codepletion. Taken together, our results identify a previously unrecognized role for MYPT1 in regulating mitosis by antagonizing PLK1.

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Local actin-dependent endocytosis is zygotically controlled to initiate Drosophila cellularization.

Publication Date: 2008 May PMID: 18477459
Authors: Sokac, A. M. - Wieschaus, E.
Journal: Dev Cell

In early Drosophila embryos, several mitotic cycles proceed with aborted cytokinesis before a modified cytokinesis, called cellularization, finally divides the syncytium into individual cells. Here, we find that scission of endocytic vesicles from the plasma membrane (PM) provides a control point to regulate the furrowing events that accompany this development. At early mitotic cycles, local furrow-associated endocytosis is controlled by cell cycle progression, whereas at cellularization, which occurs in a prolonged interphase, it is controlled by expression of the zygotic gene nullo. nullo mutations impair cortical F-actin accumulation and scission of endocytic vesicles, such that membrane tubules remain tethered to the PM and deplete structural components from the furrows, precipitating furrow regression. Thus, Nullo regulates scission to restrain endocytosis of proteins essential for furrow stabilization at the onset of cellularization. We propose that developmentally regulated endocytosis can coordinate actin/PM remodeling to directly drive furrow dynamics during morphogenesis.

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Sensory signaling-dependent remodeling of olfactory cilia architecture in C. elegans.

Publication Date: 2008 May PMID: 18477458
Authors: Mukhopadhyay, S. - Lu, Y. - Shaham, S. - Sengupta, P.
Journal: Dev Cell

Nonmotile primary cilia are sensory organelles composed of a microtubular axoneme and a surrounding membrane sheath that houses signaling molecules. Optimal cellular function requires the precise regulation of axoneme assembly, membrane biogenesis, and signaling protein targeting and localization via as yet poorly understood mechanisms. Here, we show that sensory signaling is required to maintain the architecture of the specialized AWB olfactory neuron cilia in C. elegans. Decreased sensory signaling results in alteration of axoneme length and expansion of a membraneous structure, thereby altering the topological distribution of a subset of ciliary transmembrane signaling molecules. Signaling-regulated alteration of ciliary structures can be bypassed by modulation of intracellular cGMP or calcium levels and requires kinesin-II-driven intraflagellar transport (IFT), as well as BBS- and RAB8-related proteins. Our results suggest that compensatory mechanisms in response to altered levels of sensory activity modulate AWB cilia architecture, revealing remarkable plasticity in the regulation of cilia structure.

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The C. elegans SYS-1 protein is a bona fide beta-catenin.

Publication Date: 2008 May PMID: 18477457
Authors: Liu, J. - Phillips, B. T. - Amaya, M. F. - Kimble, J. - Xu, W.
Journal: Dev Cell

C. elegans SYS-1 has key functional characteristics of a canonical beta-catenin, but no significant sequence similarity. Here, we report the SYS-1 crystal structure, both on its own and in a complex with POP-1, the C. elegans TCF homolog. The two structures possess signature features of canonical beta-catenin and the beta-catenin/TCF complex that could not be predicted by sequence. Most importantly, SYS-1 bears 12 armadillo repeats and the SYS-1/POP-1 interface is anchored by a conserved salt-bridge, the "charged button." We also modeled structures for three other C. elegans beta-catenins to predict the molecular basis of their distinct binding properties. Finally, we generated a phylogenetic tree, using the region of highest structural similarity between SYS-1 and beta-catenin, and found that SYS-1 clusters robustly within the beta-catenin clade. We conclude that the SYS-1 protein belongs to the beta-catenin family and suggest that additional divergent beta-catenins await discovery.

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Crystal structure analysis reveals how the Chordin family member crossveinless 2 blocks BMP-2 receptor binding.

Publication Date: 2008 May PMID: 18477456
Authors: Zhang, J. L. - Qiu, L. Y. - Kotzsch, A. - Weidauer, S. - Patterson, L. - Hammerschmidt, M. - Sebald, W. - Mueller, T. D.
Journal: Dev Cell

Crossveinless 2 (CV-2) is an extracellular BMP modulator protein belonging to the Chordin family. During development it is expressed at sites of high BMP signaling and like Chordin CV-2 can either enhance or inhibit BMP activity. CV-2 binds to BMP-2 via its N-terminal Von Willebrand factor type C (VWC) domain 1. Here we report the structure of the complex between CV-2 VWC1 and BMP-2. The tripartite VWC1 binds BMP-2 only through a short N-terminal segment, called clip, and subdomain (SD) 1. Mutational analysis establishes that the clip segment and SD1 together create high-affinity BMP-2 binding. All four receptor-binding sites of BMP-2 are blocked in the complex, demonstrating that VWC1 acts as competitive inhibitor for all receptor types. In vivo experiments reveal that the BMP-enhancing (pro-BMP) activity of CV-2 is independent of BMP-2 binding by VWC1, showing that pro- and anti-BMP activities are structurally separated in CV-2.

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Feedback inhibition of Jak/STAT signaling by apontic is required to limit an invasive cell population.

Publication Date: 2008 May PMID: 18477455
Authors: Starz-Gaiano, M. - Melani, M. - Wang, X. - Meinhardt, H. - Montell, D. J.
Journal: Dev Cell

In both normal development and in a variety of pathological conditions, epithelial cells can acquire migratory and invasive properties. Border cells in the Drosophila ovary provide a genetically tractable model for elucidating the mechanisms controlling such behaviors. Here we report the identification of a mutant, apontic (apt), in which the migratory population expanded and separation from the epithelium was impeded. This phenotype resembled gain-of-function of JAK/STAT activity. Gain-of-function of APT also mimicked loss of function of STAT and its key downstream target, SLBO. APT expression was induced by STAT, which bound directly to sites in the apt gene. The data suggest that a regulatory circuit between STAT, APT, and SLBO functions to convert an initially graded signal into an all-or-nothing activation of JAK/STAT and thus to proper cell specification and migration. These findings are supported by a mathematical model, which accurately simulates wild-type and mutant phenotypes.

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Cellular trafficking of the glypican Dally-like is required for full-strength Hedgehog signaling and wingless transcytosis.

Publication Date: 2008 May PMID: 18477454
Authors: Gallet, A. - Staccini-Lavenant, L. - Therond, P. P.
Journal: Dev Cell

Hedgehog (Hh) and Wingless (Wg) morphogens specify cell fate in a concentration-dependent manner in the Drosophila wing imaginal disc. Proteoglycans, components of the extracellular matrix, are involved in Hh and Wg stability, spreading, and reception. In this study, we demonstrate that the glycosyl-phosphatidyl-inositol (GPI) anchor of the glypican Dally-like (Dlp) is required for its apical internalization and its subsequent targeting to the basolateral compartment of the epithelium. Dlp endocytosis from the apical surface of Hh-receiving cells catalyzes the internalization of Hh bound to its receptor Patched (Ptc). The cointernalization of Dlp with the Hh/Ptc complex is dynamin dependent and necessary for full-strength Hh signaling. We also demonstrate that Wg is secreted apically in the disc epithelium and that apicobasal trafficking of Dlp allows Wg transcytosis to favor Wg spreading along the basolateral compartment. Thus, Dlp endocytosis is a common regulatory mechanism of both Hh and Wg morphogen action.

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Glypican-3 inhibits Hedgehog signaling during development by competing with patched for Hedgehog binding.

Publication Date: 2008 May PMID: 18477453
Authors: Capurro, M. I. - Xu, P. - Shi, W. - Li, F. - Jia, A. - Filmus, J.
Journal: Dev Cell

Loss-of-function mutations in glypican-3 (GPC3), one of the six mammalian glypicans, causes the Simpson-Golabi-Behmel overgrowth syndrome (SGBS), and GPC3 null mice display developmental overgrowth. Because the Hedgehog signaling pathway positively regulates body size, we hypothesized that GPC3 acts as an inhibitor of Hedgehog activity during development. Here, we show that GPC3 null embryos display increased Hedgehog signaling and that GPC3 inhibits Hedgehog activity in cultured mouse embryonic fibroblasts. In addition, we report that GPC3 interacts with high affinity with Hedgehog but not with its receptor, Patched, and that GPC3 competes with Patched for Hedgehog binding. Furthermore, GPC3 induces Hedgehog endocytosis and degradation. Surprisingly, the heparan sulfate chains of GPC3 are not required for its interaction with Hedgehog. We conclude that GPC3 acts as a negative regulator of Hedgehog signaling during mammalian development and that the overgrowth observed in SGBS patients is, at least in part, the consequence of hyperactivation of the Hedgehog signaling pathway.

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Patched1 haploinsufficiency increases adult bone mass and modulates Gli3 repressor activity.

Publication Date: 2008 May PMID: 18477452
Authors: Ohba, S. - Kawaguchi, H. - Kugimiya, F. - Ogasawara, T. - Kawamura, N. - Saito, T. - Ikeda, T. - Fujii, K. - Miyajima, T. - Kuramochi, A. - Miyashita, T. - Oda, H. - Nakamura, K. - Takato, T. - Chung, U. I.
Journal: Dev Cell

Hedgehog (Hh)-Patched1 (Ptch1) signaling plays essential roles in various developmental processes, but little is known about its role in postnatal homeostasis. Here, we demonstrate regulation of postnatal bone homeostasis by Hh-Ptch1 signaling. Ptch1-deficient (Ptch1+/-) mice and patients with nevoid basal cell carcinoma syndrome showed high bone mass in adults. In culture, Ptch1+/- cells showed accelerated osteoblast differentiation, enhanced responsiveness to the runt-related transcription factor 2 (Runx2), and reduced generation of the repressor form of Gli3 (Gli3rep). Gli3rep inhibited DNA binding by Runx2 in vitro, suggesting a mechanism that could contribute to the bone phenotypes seen in the Ptch1 heterozygotes. Moreover, systemic administration of the Hh signaling inhibitor cyclopamine decreased bone mass in adult mice. These data provide evidence that Hh-Ptch1 signaling plays a crucial role in postnatal bone homeostasis and point to Hh-Ptch1 signaling as a potential molecular target for the treatment of osteoporosis.

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Hedgehog signaling in mature osteoblasts regulates bone formation and resorption by controlling PTHrP and RANKL expression.

Publication Date: 2008 May PMID: 18477451
Authors: Mak, K. K. - Bi, Y. - Wan, C. - Chuang, P. T. - Clemens, T. - Young, M. - Yang, Y.
Journal: Dev Cell

Hedgehog (Hh) signaling is required for osteoblast differentiation from mesenchymal progenitors during endochondral bone formation. However, the role of Hh signaling in differentiated osteoblasts during adult bone homeostasis remains to be elucidated. We found that in the postnatal bone, Hh signaling activity was progressively reduced as osteoblasts mature. Upregulating Hh signaling selectively in mature osteoblasts led to increased bone formation and excessive bone resorption. As a consequence, these mutant mice showed severe osteopenia. Conversely, inhibition of Hh signaling in mature osteoblasts resulted in increased bone mass and protection from bone loss in older mice. Cellular and molecular studies showed that Hh signaling indirectly induced osteoclast differentiation by upregulating osteoblast expression of PTHrP, which promoted RANKL expression via PKA and its target transcription factor CREB. Our results demonstrate that Hh signaling in mature osteoblasts regulates both bone formation and resorption and that inhibition of Hh signaling reduces bone loss in aged mice.

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Glucose restriction inhibits skeletal myoblast differentiation by activating SIRT1 through AMPK-mediated regulation of Nampt.

Publication Date: 2008 May PMID: 18477450
Authors: Fulco, M. - Cen, Y. - Zhao, P. - Hoffman, E. P. - McBurney, M. W. - Sauve, A. A. - Sartorelli, V.
Journal: Dev Cell

It is intuitive to speculate that nutrient availability may influence differentiation of mammalian cells. Nonetheless, a comprehensive complement of the molecular determinants involved in this process has not been elucidated yet. Here, we have investigated how nutrients (glucose) affect skeletal myogenesis. Glucose restriction (GR) impaired differentiation of skeletal myoblasts and was associated with activation of the AMP-activated protein kinase (AMPK). Activated AMPK was required to promote GR-induced transcription of the NAD+ biosynthetic enzyme Nampt. Indeed, GR augmented the Nampt activity, which consequently modified the intracellular [NAD+]:[NADH] ratio and nicotinamide levels, and mediated inhibition of skeletal myogenesis. Skeletal myoblasts derived from SIRT1+/- heterozygous mice were resistant to the effects of either GR or AMPK activation. These experiments reveal that AMPK, Nampt, and SIRT1 are the molecular components of a functional signaling pathway that allows skeletal muscle cells to sense and react to nutrient availability.

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Polo on the Rise-from Mitotic Entry to Cytokinesis with Plk1.

Publication Date: 2008 May PMID: 18477449
Authors: Petronczki, M. - Lenart, P. - Peters, J. M.
Journal: Dev Cell

Polo-like kinase 1 (Plk1) is a key regulator of cell division in eukaryotic cells. New techniques, including the application of small-molecule inhibitors, have greatly expanded our knowledge of the functions, targets, and regulation of this key mitotic enzyme. In this review, we focus on how Plk1 is recruited to centrosomes, kinetochores, and the spindle midzone and what the specific tasks of Plk1 at these distinct subcellular structures might be. In particular, we highlight new work on the role of Plk1 in cytokinesis in human cells. Finally, we describe how better understanding of Plk1 functions allows critical evaluation of Plk1 as a potential drug target for cancer therapy.

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Cyclins in meiosis: lost in translation.

Publication Date: 2008 May PMID: 18477448
Authors: Futcher, B.
Journal: Dev Cell

In a recent issue of Cell, Carlile and Amon examine the regulation of four budding yeast B-type cyclins, crucial for regulating and distinguishing meoisis I and meoisis II divisions, and find a surprising diversity of behaviors and modes of regulation. In particular, Clb3 is regulated by a striking translational repression specific to meoisis I.

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Glucose restriction: longevity SIRTainly, but without building muscle?

Publication Date: 2008 May PMID: 18477447
Authors: Canto, C. - Auwerx, J.
Journal: Dev Cell

The two metabolic sensors AMPK and SIRT1 take center stage as Fulco et al. reveal, in this issue of Developmental Cell, the signaling mechanism by which low glucose prevents the correct development of the myogenic program. These observations may hold some therapeutic promise against muscle wasting.

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Together we stand: genes cluster to coordinate regulation.

Publication Date: 2008 May PMID: 18477446
Authors: Amoutzias, G. - Van de Peer, Y.
Journal: Dev Cell

Although most eukaryotic genomes lack operons, occasionally clusters of genes are discovered that are related in function. Now, a metabolic operon-like gene cluster has been described in Arabidopsis thaliana, that is needed for triterpene synthesis.

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A topographical map of spatiotemporal patterns of gene expression.

Publication Date: 2008 May PMID: 18477445
Authors: Furlong, E. E.
Journal: Dev Cell

A recent study by Folkes et al. in Cell generated a 3D atlas of gene expression for the Drosophila blastoderm embryo using a new approach for image registration. This virtual embryo allows in silico multiplexing of in situ hybridizations and lays the groundwork for new insights into gene regulatory networks.

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Hedgehog coordination of postnatal osteoclast and osteoblast activities.

Publication Date: 2008 May PMID: 18477444
Authors: Mundy, G. R. - Yang, X.
Journal: Dev Cell

The Hedgehog (Hh) pathway is important for skeletal patterning and morphogenesis during embryonic development. Papers by Ohba et al. and Mak et al. in this edition of Developmental Cell suggest that Hh signaling may exert delicate control over the activities of osteoclasts and osteoblasts, the cell types primarily responsible for bone resorption and formation.

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A cilium is not a cilium is not a cilium: signaling contributes to ciliary morphological diversity.

Publication Date: 2008 May PMID: 18477443
Authors: Reiter, J. F.
Journal: Dev Cell

Mukhopadhyay and colleagues reveal in this issue of Developmental Cell that signaling mediated by a specialized neuronal cilium in C. elegans affects its structure. The finding that this cilium is modified in response to the cues it transduces suggests that cilia may not be static antennae, but organelles whose functions are shaped by their signaling activities.

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