Hot off the presses! Jan 18 dev cell

The Jan 18 issue of the dev cell is now up on Pubget (About dev cell): if you're at a subscribing institution, just click the link in the latest link at the home page. (Note you'll only be able to get all the PDFs in the issue if your institution subscribes to Pubget.)

Latest Articles Include:

• Introducing ESCRTs II and III
  - dev cell 20(1):e1 (2011)
  As someone who likes solving the molecular puzzle of protein complex assembly, I enjoyed reading two back-to-back articles by Scott Emr and colleagues describing the ESCRT-II and ESCRT-III components of the multivesicular body (MVB)-sorting machinery. Not only was the biochemical evidence top notch, it was also firmly grounded on the genetics of the system. These studies, together with the previous identification of ESCRT-I by the same group published in Cell a year earlier, set the basis for all current analyses of MVB formation and function. Thanks to this work, what was once just a tantalizing morphological entity took center stage in studies of the targeting of transmembrane proteins for degradation in lysosomes. Moreover, the ability of the ESCRT complexes to sculpt intraluminal vesicles from the limiting membrane of endosomes turned out to mediate other important biological processes such as membrane scission during cytokinesis and the release of enveloped viruse! s, thus becoming a paradigm for membrane budding away from the cytosol. This PaperPick refers to "Escrt-III: An Endosome-Associated Hetero-oligomeric Protein Complex Required for MVB Sorting" by M. Babst, D.J. Katzmann, E.J. Estepa-Sabal, T. Meerloo, and S.D. Emr and "Endosome-Associated Complex, ESCRT-II, Recruits Transport Machinery for Protein Sorting at the Multivesicular Body" by M. Babst, D.J. Katzmann, W.B. Snyder, B. Wendland, and S.D. Emr, both published in August, 2002. Finding ESCRTs II and III (24733 K) Markus Babst and David Katzmann talk about how the ESCRT project started and developed in the Emr lab and where ESCRTs have gone from there.
• Finding Centralspindlin
  - dev cell 20(1):e2 (2011)
  In this paper, Mishima, Kaitna, and Glotzer reported the discovery and characterization of a widely conserved tetrameric protein complex that they dubbed centralspindlin. Using an impressive combination of in vitro biochemistry and in vivo genetics, and including work in both C. elegans and mammalian cell culture, they showed that centralspindlin functions as a complex—not just as individual components—and is required for assembly of the central mitotic spindle and for cytokinesis. We also learned that centralspindlin contains two copies each of a RhoGAP and a kinesin 6 family member (CYK-4 and ZEN-4 in C. elegans; MgcRacGAP and MKLP1 in vertebrates), where the protein-protein interaction domains lie, and that centralspindlin bundles microtubules in vitro. This paper firmly established C. elegans as an important model system for investigating the fundamental cellular process of cytokinesis and laid the foundation for many subsequent studies that have provided subst! antial mechanistic insight into how the mitotic spindle interacts with and regulates furrow formation during cytokinesis. This PaperPick refers to "Central Spindle Assembly and Cytokinesis Require a Kinesin-like Protein/RhoGAP Complex with Microtubule Bundling Activity" by M. Mishima, S. Kaitna, and M. Glotzer, published in January 2002. The Ins and Outs of Centralspindlin (10566 K) Michael Glotzer takes us on a personal tour of the paper from his group that first described centralspindlin and discusses how our understanding of this complex has developed from there.
• Going on a (Membrane) Bender to the F-BAR
  - dev cell 20(1):e3 (2011)
  This study is a personal favorite because it simplified the field and proposed an elegant model of how actin and membranes work together in endocytic trafficking. A group of proteins containing FCH domains (Fes/CIP4 homology) were discovered to have extended homology with the banana-shaped alpha helical BAR domain and renamed F-BAR proteins. F-BAR and BAR proteins bind to and bend curved membrane surfaces and participate in budding and tubulation events. Dynamin, a large GTPase that forms helical cages around tubulated membranes, is the main partner of BAR and F-BAR proteins. The actin cytoskeleton opposes scission of the tubulated membranes pending arrival of an appropriate signal. The idea that dynamin is the most important partner of F-BARs and that the trio of actin, F-BAR, and dynamin sculpt and cut membrane tubules in this elegant way remains important for future research and puts into context the barrage of sometimes disparate studies on these proteins. This PaperPick refers to "Dynamin and the Actin Cytoskeleton Cooperatively Regulate Plasma Membrane Invagination by BAR and F-BAR Proteins" by T. Itoh, K.S. Erdmann, A. Roux, B. Habermann, H. Werner, and P. De Camilli, published in December, 2005. F-BAR Proteins: Coordinating the Effects of Actin and Dynamin on Endocytosis (11857 K) Pietro De Camilli discusses how his group came to define the F-BAR domain, its role in endocytosis, and where subsequent work has taken the field.
• Shedding Light on Mammalian Microautophagy
  - dev cell 20(1):1-2 (2011)
  ESCRT complexes are implicated in mediating membrane protein degradation, whereas hsc70 mediates cytosolic protein degradation via chaperone-mediated autophagy. In this issue of Developmental Cell, Sahu et al. (2011) describe in mammalian cells the involvement of ESCRT complexes and hsc70 in the degradation of cytosolic proteins in a process resembling microautophagy.
• MicroRNAs Sculpt Gene Expression in Embryonic Development: New Insights from Plants
  - dev cell 20(1):3-4 (2011)
  Zygotic microRNAs coordinate the clearance of maternal mRNA in animals to facilitate developmental transitions. In a recent issue of Genes and Development, Nodine and Bartel (2010) uncover a reciprocal function in plants, where miRNA-156 preemptively represses genes that function later in development to prevent premature developmental transitions.
• Epigenetic Memory Meets G2/M: To Remember or To Forget?
  - dev cell 20(1):5-6 (2011)
  The histone H3 lysine 27 (H3K27) methyltransferase EZH2 is essential for stem cell maintenance and proliferation. Recent insights suggest that the cyclin-dependent kinase CDK1 phosphorylates EZH2 at specific threonine residues by sensing developmental cues to mediate self-renewal or differentiation during G2/M phase.
• A Receptor for Meningococcus: Eliciting β-Arrestin Signaling for Barrier Breaching
  - dev cell 20(1):7-8 (2011)
  In a recent issue of Cell, Coureuil et al. (2010) show that type IV pili of Neisseria meningitidis—the causative bacterium of cerebrospinal meningitis—bind to the endothelial β2-adrenoceptor and elicit biased β-arrestin signaling resulting in microvilli formation and paracytotic bacterial migration.
• Whence Directionality: Guidance Mechanisms in Solitary and Collective Cell Migration
  - dev cell 20(1):9-18 (2011)
  As individual cells or groups of cells move through the complex environment of the body, their migration is affected by multiple external cues. Some cues are diffusible signaling molecules, and some are solid biophysical features. How do cells respond appropriately? This perspective discusses the relationship between guidance input and the cellular output, considering effects from classical chemotaxis to contact-dependent guidance. The influences of membrane trafficking and of imposed constraints on directional movement are also considered. New insights regarding guidance and dynamic cell polarity have emerged from examining new cell migration models and from re-examining well known ones with new approaches and new tools.
• MicroRNA-9 Reveals Regional Diversity of Neural Progenitors along the Anterior-Posterior Axis
  - dev cell 20(1):19-32 (2011)
  Neural progenitors self-renew and generate neurons throughout the central nervous system. Here, we uncover an unexpected regional specificity in the properties of neural progenitor cells, revealed by the function of a microRNA—miR-9. miR-9 is expressed in neural progenitors, and its knockdown results in an inhibition of neurogenesis along the anterior-posterior axis. However, the underlying mechanism differs—in the hindbrain, progenitors fail to exit the cell cycle, whereas in the forebrain they undergo apoptosis, counteracting the proliferative effect. Among several targets, we functionally identify hairy1 as a primary target of miR-9, regulated at the mRNA level. hairy1 mediates the effects of miR-9 on proliferation, through Fgf8 signaling in the forebrain and Wnt signaling in the hindbrain, but affects apoptosis only in the forebrain, via the p53 pathway. Our findings show a positional difference in the responsiveness of progenitors to miR-9 depletion, revealing! an underlying divergence of their properties.
• Robo4 Maintains Vessel Integrity and Inhibits Angiogenesis by Interacting with UNC5B
  - dev cell 20(1):33-46 (2011)
  Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To identify Robo4 binding partners, we performed a protein-protein interaction screen with the Robo4 extracellular domain. We find that Robo4 specifically binds to UNC5B, a vascular Netrin receptor, revealing unexpected interactions between two endothelial guidance receptors. We show that Robo4 maintains vessel integrity by activating UNC5B, which inhibits signaling downstream of vascular endothelial growth factor (VEGF). Function-blocking monoclonal antibodies against Robo4 and UNC5B increase angiogenesis and disrupt vessel integrity. Soluble Robo4 protein inhibits VEGF-induced vessel permeability and rescues barrier defects in Robo4−/− mice, but not in mice treated with anti-UNC5B. Thus, Robo4-UNC5B signaling maintains vascular integrity by counteracting VEGF signaling in endothelial cells, identifying a novel function of guidance receptor interactions in the vasculature.
• PI4P and Rab Inputs Collaborate in Myosin-V-Dependent Transport of Secretory Compartments in Yeast
  - dev cell 20(1):47-59 (2011)
  Cell polarity involves transport of specific membranes and macromolecules at the right time to the right place. In budding yeast, secretory vesicles are transported by the myosin-V Myo2p to sites of cell growth. We show that phosphatidylinositol 4-phosphate (PI4P) is present in late secretory compartments and is critical for their association with, and transport by, Myo2p. Further, the trans-Golgi network Rab Ypt31/32p and secretory vesicle Rab Sec4p each bind directly, but distinctly, to Myo2p, and these interactions are also required for secretory compartment transport. Enhancing the interaction of Myo2p with PI4P bypasses the requirement for interaction with Ypt31/32p and Sec4p. Together with additional genetic data, the results indicate that Rab proteins and PI4P collaborate in the association of secretory compartments with Myo2p. Thus, we show that a coincidence detection mechanism coordinates inputs from PI4P and the appropriate Rab for secretory compartment tran! sport.
• KIF16B/Rab14 Molecular Motor Complex Is Critical for Early Embryonic Development by Transporting FGF Receptor
  - dev cell 20(1):60-71 (2011)
  Kinesin-mediated membrane trafficking is a fundamental cellular process, but its developmental relevance is little understood. Here we show that the kinesin-3 motor KIF16B/Rab14 complex acts in biosynthetic Golgi-to-endosome traffic of the fibroblast growth factor receptor (FGFR) during early embryonic development. Kif16b–/– mouse embryos failed in developing epiblast and primitive endoderm lineages and died in the peri-implantation stage, similar to previously reported FGFR2 knockout embryos. KIF16B associated directly with the Rab14-GTP adaptor on FGFR-containing vesicles and transported them toward the plasma membrane. To examine whether the nucleotide state of Rab14 serves as a switch for transport, we performed Rab14-GDP overexpression. This dominant negative approach reproduced the whole putative sequence of KIF16B or FGFR2 deficiency: impairment in FGFR transport, FGF signaling, basement membrane assembly by the primitive endoderm lineage, and epiblast devel! opment. These data provide one of the first pieces of genetic evidence that microtubule-based membrane trafficking directly promotes early development.
• Brat Promotes Stem Cell Differentiation via Control of a Bistable Switch that Restricts BMP Signaling
  - dev cell 20(1):72-83 (2011)
  Drosophila ovarian germline stem cells (GSCs) are maintained by Dpp signaling and the Pumilio (Pum) and Nanos (Nos) translational repressors. Upon division, Dpp signaling is extinguished, and Nos is downregulated in one daughter cell, causing it to switch to a differentiating cystoblast (CB). However, downstream effectors of Pum-Nos remain unknown, and how CBs lose their responsiveness to Dpp is unclear. Here, we identify Brain Tumor (Brat) as a potent differentiation factor and target of Pum-Nos regulation. Brat is excluded from GSCs by Pum-Nos but functions with Pum in CBs to translationally repress distinct targets, including the Mad and dMyc mRNAs. Regulation of both targets simultaneously lowers cellular responsiveness to Dpp signaling, forcing the cell to become refractory to the self-renewal signal. Mathematical modeling elucidates bistability of cell fate in the Brat-mediated system, revealing how autoregulation of GSC number can arise from Brat coupling extrac! ellular Dpp regulation to intracellular interpretation.
• Ras Effector Switching Promotes Divergent Cell Fates in C. elegans Vulval Patterning
  - dev cell 20(1):84-96 (2011)
  The C. elegans vulva is patterned by epidermal growth factor (EGF) activation of Ras to control 1° fate, and 1° fate induces antagonistic Notch-dependent 2° fate. Furthermore, a spatial EGF gradient, in addition to inducing 1° fate, directly contributes to 2° fate via an unknown pathway. We find that in addition to its canonical effector, Raf, vulval Ras utilizes an exchange factor for the Ral small GTPase (RalGEF), such that Ras-RalGEF-Ral antagonizes Ras-Raf pro-1° fate activity. Consistent with its restricted expression pattern, Ral participates in EGF pro-2° activity. Thus, we have delineated a Ras effector-switching mechanism whereby position within the morphogen gradient dictates that Ras effector usage is switched to RalGEF from Raf to promote 2° instead of 1° fate. Our observations define the utility of Ras effector switching during normal development and may provide a possible mechanistic basis for cell and cancer-type differences in effector dependen! cy and activation.
• Rheb1 Is Required for mTORC1 and Myelination in Postnatal Brain Development
  - dev cell 20(1):97-108 (2011)
  mTor kinase is involved in cell growth, proliferation, and differentiation. The roles of mTor activators, Rheb1 and Rheb2, have not been established in vivo. Here, we report that Rheb1, but not Rheb2, is critical for embryonic survival and mTORC1 signaling. Embryonic deletion of Rheb1 in neural progenitor cells abolishes mTORC1 signaling in developing brain and increases mTORC2 signaling. Remarkably, embryonic and early postnatal brain development appears grossly normal in these Rheb1f/f,Nes-cre mice with the notable exception of deficits of myelination. Conditional expression of Rheb1 transgene in neural progenitors increases mTORC1 activity and promotes myelination in the brain. In addition the Rheb1 transgene rescues mTORC1 signaling and hypomyelination in the Rheb1f/f,Nes-cre mice. Our study demonstrates that Rheb1 is essential for mTORC1 signaling and myelination in the brain, and suggests that mTORC1 signaling plays a role in selective cellular adaptations, rathe! r than general cellular viability.
• Cooperative Regulation of Growth by Yorkie and Mad through bantam
  - dev cell 20(1):109-122 (2011)
  The Dpp and Fat-Hippo signaling pathways both regulate growth in Drosophila. Dpp is a BMP family ligand and acts via a Smad family DNA-binding transcription factor, Mad. Fat-Hippo signaling acts via a non-DNA-binding transcriptional coactivator protein, Yorkie. Here, we show that these pathways are directly interlinked. They act synergistically to promote growth, in part via regulation of the microRNA gene bantam, and their ability to promote growth is mutually dependent. Yorkie and Mad physically bind each other, and we identify a 410 bp minimal enhancer of bantam that responds to Yorkie:Mad in vivo and in cultured cells, and show that both Yorkie and Mad associate with this enhancer in vivo. Our results indicate that in promoting the growth of Drosophila tissues, Fat-Hippo and Dpp signaling contribute distinct subunits of a shared transcriptional activation complex, Yorkie:Mad.
• Antagonistic Growth Regulation by Dpp and Fat Drives Uniform Cell Proliferation
  - dev cell 20(1):123-130 (2011)
  We use the Dpp morphogen gradient in the Drosophila wing disc as a model to address the fundamental question of how a gradient of a growth factor can produce uniform growth. We first show that proper expression and subcellular localization of components in the Fat tumor-suppressor pathway, which have been argued to depend on Dpp activity differences, are not reliant on the Dpp gradient. We next analyzed cell proliferation in discs with uniformly high Dpp or uniformly low Fat signaling activity and found that these pathways regulate growth in a complementary manner. While the Dpp mediator Brinker inhibits growth in the primordium primarily in the lateral regions, Fat represses growth mostly in the medial region. Together, our results indicate that the activities of both signaling pathways are regulated in a parallel rather than sequential manner and that uniform proliferation is achieved by their complementary action on growth.
• Microautophagy of Cytosolic Proteins by Late Endosomes
  - dev cell 20(1):131-139 (2011)
  Autophagy delivers cytosolic components to lysosomes for their degradation. The delivery of autophagic cargo to late endosomes for complete or partial degradation has also been described. In this report we present evidence that distinct autophagic mechanisms control cytosolic protein delivery to late endosomes and identify a microautophagy-like process that delivers soluble cytosolic proteins to the vesicles of late endosomes/multivesicular bodies (MVBs). This microautophagy-like process has selectivity and is distinct from chaperone-mediated autophagy that occurs in lysosomes. Endosomal microautophagy occurs during MVB formation, relying on the ESCRT I and III systems for formation of the vesicles in which the cytosolic cargo is internalized. Protein cargo selection is mediated by the chaperone hsc70 and requires the cationic domain of hsc70 for electrostatic interactions with the endosomal membrane. Therefore, we propose that endosomal microautophagy shares molecular! components with both the endocytic and autophagic pathways.