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Waking up muscle stem cells: PI3K signalling is ringing
Adult skeletal muscle injury instructs endogenous quiescent stem cells to activate for repair. The pathways involved in this activation remain poorly understood. An elegant study by Wang et al (2018) combine mouse molecular genetics with muscle regeneration experiments, cell culture and transcriptomic analysis to show that early activation depends on a PI3K-mTORC1-Jun/FoxOs signalling axis. (Source: EMBO Journal)
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Relaix, F., Machado, L. Tags: Cell Cycle, Signal Transduction, Stem Cells News [amp ] Views Source Type: research

Fat nerves keep pain at bay
Inflammatory pain is a debilitating condition and a severe health burden; physiologically, it is a complex phenomenon with multiple contributing mechanisms. A new study published in The EMBO Journal reports that tissue inflammation results in local depletion of cholesterol in nociceptive nerves, causing a loss of lipid raft localization of a sodium channel Nav1.9 and, ultimately, resulting in potentiation of its activity. The discovered effect contributes to the inflammatory overexcitability of peripheral nociceptive nerve terminals resulting in inflammatory hyperalgesia. Topical application of cholesterol-containing ...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Shah, S., Gamper, N. Tags: Immunology, Neuroscience News [amp ] Views Source Type: research

p110{alpha} of PI3K is necessary and sufficient for quiescence exit in adult muscle satellite cells
Adult mouse muscle satellite cells (MuSCs) are quiescent in uninjured muscles. Upon injury, MuSCs exit quiescence in vivo to become activated, re-enter the cell cycle to proliferate, and differentiate to repair the damaged muscles. It remains unclear which extrinsic cues and intrinsic signaling pathways regulate quiescence exit during MuSC activation. Here, we demonstrated that inducible MuSC-specific deletion of p110α, a catalytic subunit of phosphatidylinositol 3-kinase (PI3K), rendered MuSCs unable to exit quiescence, resulting in severely impaired MuSC proliferation and muscle regeneration. Genetic reactivat...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Wang, G., Zhu, H., Situ, C., Han, L., Yu, Y., Cheung, T. H., Liu, K., Wu, Z. Tags: Cell Cycle, Signal Transduction, Stem Cells Articles Source Type: research

OST1-mediated BTF3L phosphorylation positively regulates CBFs during plant cold responses
In this study, we identified BTF3 and BTF3L (BTF3-like), β-subunits of a nascent polypeptide-associated complex (NAC), as OST1 substrates that positively regulate freezing tolerance. OST1 phosphorylates BTF3 and BTF3L in vitro and in vivo, and facilitates their interaction with C-repeat-binding factors (CBFs) to promote CBF stability under cold stress. The phosphorylation of BTF3L at the Ser50 residue by OST1 is required for its function in regulating freezing tolerance. In addition, BTF3 and BTF3L proteins positively regulate the expression of CBF genes. These findings unravel a molecular mechanism by which...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Ding, Y., Jia, Y., Shi, Y., Zhang, X., Song, C., Gong, Z., Yang, S. Tags: Physiology, Plant Biology, Signal Transduction Articles Source Type: research

Regulation of RNA polymerase II processivity by Spt5 is restricted to a narrow window during elongation
Spt5 is a highly conserved RNA polymerase II (Pol II)-associated pausing and elongation factor. However, its impact on global elongation and Pol II processivity in mammalian cells has not been clarified. Here, we show that depleting Spt5 in mouse embryonic fibroblasts (MEFs) does not cause global elongation defects or decreased elongation rates. Instead, in Spt5-depleted cells, a fraction of Pol II molecules are dislodged during elongation, thus decreasing the number of Pol II complexes that complete the transcription cycle. Most strikingly, this decrease is restricted to a narrow window between 15 and 20 kb from the ...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Fitz, J., Neumann, T., Pavri, R. Tags: Chromatin, Epigenetics, Genomics & Functional Genomics, Transcription Articles Source Type: research

Mitotic spindle association of TACC3 requires Aurora-A-dependent stabilization of a cryptic {alpha}-helix
Aurora-A regulates the recruitment of TACC3 to the mitotic spindle through a phospho-dependent interaction with clathrin heavy chain (CHC). Here, we describe the structural basis of these interactions, mediated by three motifs in a disordered region of TACC3. A hydrophobic docking motif binds to a previously uncharacterized pocket on Aurora-A that is blocked in most kinases. Abrogation of the docking motif causes a delay in late mitosis, consistent with the cellular distribution of Aurora-A complexes. Phosphorylation of Ser558 engages a conformational switch in a second motif from a disordered state, needed to bind the kin...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Burgess, S. G., Mukherjee, M., Sabir, S., Joseph, N., Gutierrez-Caballero, C., Richards, M. W., Huguenin-Dezot, N., Chin, J. W., Kennedy, E. J., Pfuhl, M., Royle, S. J., Gergely, F., Bayliss, R. Tags: Cell Adhesion, Polarity & Cytoskeleton, Cell Cycle, Structural Biology Articles Source Type: research

Attenuation of cGAS-STING signaling is mediated by a p62/SQSTM1-dependent autophagy pathway activated by TBK1
Negative regulation of immune pathways is essential to achieve resolution of immune responses and to avoid excess inflammation. DNA stimulates type I IFN expression through the DNA sensor cGAS, the second messenger cGAMP, and the adaptor molecule STING. Here, we report that STING degradation following activation of the pathway occurs through autophagy and is mediated by p62/SQSTM1, which is phosphorylated by TBK1 to direct ubiquitinated STING to autophagosomes. Degradation of STING was impaired in p62-deficient cells, which responded with elevated IFN production to foreign DNA and DNA pathogens. In the absence of p62, STIN...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Prabakaran, T., Bodda, C., Krapp, C., Zhang, B.-c., Christensen, M. H., Sun, C., Reinert, L., Cai, Y., Jensen, S. B., Skouboe, M. K., Nyengaard, J. R., Thompson, C. B., Lebbink, R. J., Sen, G. C., van Loo, G., Nielsen, R., Komatsu, M., Nejsum, L. N., Jako Tags: Immunology, Signal Transduction Articles Source Type: research

Transmembrane redox control and proteolysis of PdeC, a novel type of c-di-GMP phosphodiesterase
The nucleotide second messenger c-di-GMP nearly ubiquitously promotes bacterial biofilm formation, with enzymes that synthesize and degrade c-di-GMP being controlled by diverse N-terminal sensor domains. Here, we describe a novel class of widely occurring c-di-GMP phosphodiesterases (PDE) that feature a periplasmic "CSS domain" with two highly conserved cysteines that is flanked by two transmembrane regions (TM1 and TM2) and followed by a cytoplasmic EAL domain with PDE activity. Using PdeC, one of the five CSS domain PDEs of Escherichia coli K-12, we show that DsbA/DsbB-promoted disulfide bond formation in the C...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Herbst, S., Lorkowski, M., Sarenko, O., Nguyen, T. K. L., Jaenicke, T., Hengge, R. Tags: Microbiology, Virology & Host Pathogen Interaction, Post-translational Modifications, Proteolysis & Proteomics, Signal Transduction Articles Source Type: research

Membrane cholesterol depletion as a trigger of Nav1.9 channel-mediated inflammatory pain
In conclusion, our data establish that membrane cholesterol is a modulator of pain transmission and shed a new light on the relationship between cholesterol homeostasis, inflammation, and pain. (Source: EMBO Journal)
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Amsalem, M., Poilbout, C., Ferracci, G., Delmas, P., Padilla, F. Tags: Immunology, Neuroscience Articles Source Type: research

LncKdm2b controls self-renewal of embryonic stem cells via activating expression of transcription factor Zbtb3
Divergent long noncoding RNAs (lncRNAs) represent a major lncRNA biotype in mouse and human genomes. The biological and molecular functions of the divergent lncRNAs remain largely unknown. Here, we show that lncKdm2b, a divergent lncRNA for Kdm2b gene, is conserved among five mammalian species and highly expressed in embryonic stem cells (ESCs) and early embryos. LncKdm2b knockout impairs ESC self-renewal and causes early embryonic lethality. LncKdm2b can activate Zbtb3 by promoting the assembly and ATPase activity of Snf2-related CREBBP activator protein (SRCAP) complex in trans. Zbtb3 potentiates the ESC self-renewal in ...
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Ye, B., Liu, B., Yang, L., Zhu, X., Zhang, D., Wu, W., Zhu, P., Wang, Y., Wang, S., Xia, P., Du, Y., Meng, S., Huang, G., Wu, J., Chen, R., Tian, Y., Fan, Z. Tags: Development & Differentiation, RNA Biology, Stem Cells Articles Source Type: research

Publication catalysis--lowering activation energy
Life Science Alliance is a new platform for and by the research community to increase the efficacy of publishing high-quality research findings. (Source: EMBO Journal)
Source: EMBO Journal - April 13, 2018 Category: Molecular Biology Authors: Pulverer, B. Tags: Editorial Source Type: research

Differentiate and switch, a tale of two heads of a lipid
Neuronal differentiation is an intricate process involving many factors and programs. One notable "curiosity" has been the observation that upon neuronal differentiation, stem cells switch the expression of their surface glycosphingolipids (GSLs) by substituting one class (the globo-series) of GSLs by another (the ganglio-series). Russo and colleagues show that there is an intricate dance between these two lipid series such that the globo products suppress neuronal differentiation via the master regulator AUTS2, which in turn suppresses the formation of the ganglio-series. These findings open the door for further...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Hannun, Y. A. Tags: Metabolism, Neuroscience News [amp ] Views Source Type: research

Paligenosis: prepare to regenerate!
Tissue injury can stimulate quiescent cells to proliferate, resulting in metaplasia, to rapidly replace damaged cells and enable regeneration. A new study describes how fully differentiated cells return to proliferation in an autophagy- and mTORC1-dependent manner. Given the striking parallels between this process in mammalian stomach and pancreas, a new term, paligenosis, is proposed for this conserved program. (Source: EMBO Journal)
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Messal, H. A., Cremona, C. A., Lan, L., Behrens, A. Tags: Cell Cycle, Autophagy & Cell Death, Development & Differentiation News [amp ] Views Source Type: research

Metastasis inside-out: dissemination of cancer cell clusters with inverted polarity
Zajac et al (2018) utilize patient tissue to discover a novel cellular mechanism for metastasis in colorectal cancer. They isolated cancer cell clusters from patient peritoneal fluid and demonstrated that these clusters invaded collectively into collagen gels, patient-derived tissues, and through the peritoneum of mice. During invasion the clusters retained an inverted epithelial architecture, functioning as multicellular metastatic seeds with inverted polarity. (Source: EMBO Journal)
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Ewald, A. J. Tags: Cancer, Cell Adhesion, Polarity & Cytoskeleton, Molecular Biology of Disease News [amp ] Views Source Type: research

Regulation of Hed1 and Rad54 binding during maturation of the meiosis-specific presynaptic complex
Most eukaryotes have two Rad51/RecA family recombinases, Rad51, which promotes recombination during mitotic double-strand break (DSB) repair, and the meiosis-specific recombinase Dmc1. During meiosis, the strand exchange activity of Rad51 is downregulated through interactions with the meiosis-specific protein Hed1, which helps ensure that strand exchange is driven by Dmc1 instead of Rad51. Hed1 acts by preventing Rad51 from interacting with Rad54, a cofactor required for promoting strand exchange during homologous recombination. However, we have a poor quantitative understanding of the regulatory interplay between these pr...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Crickard, J. B., Kaniecki, K., Kwon, Y., Sung, P., Lisby, M., Greene, E. C. Tags: DNA Replication, Repair & Recombination Articles Source Type: research

Structure of a eukaryotic cytoplasmic pre-40S ribosomal subunit
Final maturation of eukaryotic ribosomes occurs in the cytoplasm and requires the sequential removal of associated assembly factors and processing of the immature 20S pre-RNA. Using cryo-electron microscopy (cryo-EM), we have determined the structure of a yeast cytoplasmic pre-40S particle in complex with Enp1, Ltv1, Rio2, Tsr1, and Pno1 assembly factors poised to initiate final maturation. The structure reveals that the pre-rRNA adopts a highly distorted conformation of its 3' major and 3' minor domains stabilized by the binding of the assembly factors. This observation is consistent with a mechanism that involves concert...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Scaiola, A., Pena, C., Weisser, M., Böhringer, D., Leibundgut, M., Klingauf-Nerurkar, P., Gerhardy, S., Panse, V. G., Ban, N. Tags: Protein Biosynthesis & Quality Control, RNA Biology, Structural Biology Articles Source Type: research

Regenerative proliferation of differentiated cells by mTORC1-dependent paligenosis
In 1900, Adami speculated that a sequence of context-independent energetic and structural changes governed the reversion of differentiated cells to a proliferative, regenerative state. Accordingly, we show here that differentiated cells in diverse organs become proliferative via a shared program. Metaplasia-inducing injury caused both gastric chief and pancreatic acinar cells to decrease mTORC1 activity and massively upregulate lysosomes/autophagosomes; then increase damage associated metaplastic genes such as Sox9; and finally reactivate mTORC1 and re-enter the cell cycle. Blocking mTORC1 permitted autophagy and metaplast...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Willet, S. G., Lewis, M. A., Miao, Z.-F., Liu, D., Radyk, M. D., Cunningham, R. L., Burclaff, J., Sibbel, G., Lo, H.-Y. G., Blanc, V., Davidson, N. O., Wang, Z.-N., Mills, J. C. Tags: Cell Cycle, Autophagy & Cell Death, Development & Differentiation Articles Source Type: research

Two distinct conformational states define the interaction of human RAD51-ATP with single-stranded DNA
An essential mechanism for repairing DNA double-strand breaks is homologous recombination (HR). One of its core catalysts is human RAD51 (hRAD51), which assembles as a helical nucleoprotein filament on single-stranded DNA, promoting DNA-strand exchange. Here, we study the interaction of hRAD51 with single-stranded DNA using a single-molecule approach. We show that ATP-bound hRAD51 filaments can exist in two different states with different contour lengths and with a free-energy difference of ~4 kBT per hRAD51 monomer. Upon ATP hydrolysis, the filaments convert into a disassembly-competent ADP-bound configuration. In agreeme...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Brouwer, I., Moschetti, T., Candelli, A., Garcin, E. B., Modesti, M., Pellegrini, L., Wuite, G. J., Peterman, E. J. Tags: DNA Replication, Repair & Recombination, Structural Biology Articles Source Type: research

Tau protein liquid-liquid phase separation can initiate tau aggregation
The transition between soluble intrinsically disordered tau protein and aggregated tau in neurofibrillary tangles in Alzheimer's disease is unknown. Here, we propose that soluble tau species can undergo liquid–liquid phase separation (LLPS) under cellular conditions and that phase-separated tau droplets can serve as an intermediate toward tau aggregate formation. We demonstrate that phosphorylated or mutant aggregation prone recombinant tau undergoes LLPS, as does high molecular weight soluble phospho-tau isolated from human Alzheimer brain. Droplet-like tau can also be observed in neurons and other cells. We found t...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Wegmann, S., Eftekharzadeh, B., Tepper, K., Zoltowska, K. M., Bennett, R. E., Dujardin, S., Laskowski, P. R., MacKenzie, D., Kamath, T., Commins, C., Vanderburg, C., Roe, A. D., Fan, Z., Molliex, A. M., Hernandez-Vega, A., Muller, D., Hyman, A. A., Mandel Tags: Physiology, Protein Biosynthesis & Quality Control Articles Source Type: research

Modeling and resistant alleles explain the selectivity of antimalarial compound 49c towards apicomplexan aspartyl proteases
Toxoplasma gondii aspartyl protease 3 (TgASP3) phylogenetically clusters with Plasmodium falciparum Plasmepsins IX and X (PfPMIX, PfPMX). These proteases are essential for parasite survival, acting as key maturases for secreted proteins implicated in invasion and egress. A potent antimalarial peptidomimetic inhibitor (49c) originally developed against Plasmepsin II selectively targets TgASP3, PfPMIX, and PfPMX. To unravel the molecular basis for the selectivity of 49c, we constructed homology models of PfPMIX, PfPMX, and TgASP3 that were first validated by identifying the determinants of microneme and rhoptry substrate rec...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Mukherjee, B., Tessaro, F., Vahokoski, J., Kursula, I., Marq, J.-B., Scapozza, L., Soldati-Favre, D. Tags: Microbiology, Virology & Host Pathogen Interaction, Post-translational Modifications, Proteolysis & Proteomics, Structural Biology Articles Source Type: research

Molecular architecture of LSM14 interactions involved in the assembly of mRNA silencing complexes
The LSM domain-containing protein LSM14/Rap55 plays a role in mRNA decapping, translational repression, and RNA granule (P-body) assembly. How LSM14 interacts with the mRNA silencing machinery, including the eIF4E-binding protein 4E-T and the DEAD-box helicase DDX6, is poorly understood. Here we report the crystal structure of the LSM domain of LSM14 bound to a highly conserved C-terminal fragment of 4E-T. The 4E-T C-terminus forms a bi-partite motif that wraps around the N-terminal LSM domain of LSM14. We also determined the crystal structure of LSM14 bound to the C-terminal RecA-like domain of DDX6. LSM14 binds DDX6 via ...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Brandmann, T., Fakim, H., Padamsi, Z., Youn, J.-Y., Gingras, A.-C., Fabian, M. R., Jinek, M. Tags: RNA Biology, Structural Biology Articles Source Type: research

Glycosphingolipid metabolic reprogramming drives neural differentiation
Neural development is accomplished by differentiation events leading to metabolic reprogramming. Glycosphingolipid metabolism is reprogrammed during neural development with a switch from globo- to ganglio-series glycosphingolipid production. Failure to execute this glycosphingolipid switch leads to neurodevelopmental disorders in humans, indicating that glycosphingolipids are key players in this process. Nevertheless, both the molecular mechanisms that control the glycosphingolipid switch and its function in neurodevelopment are poorly understood. Here, we describe a self-contained circuit that controls glycosphingolipid r...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Russo, D., Della Ragione, F., Rizzo, R., Sugiyama, E., Scalabri, F., Hori, K., Capasso, S., Sticco, L., Fioriniello, S., De Gregorio, R., Granata, I., Guarracino, M. R., Maglione, V., Johannes, L., Bellenchi, G. C., Hoshino, M., Setou, M., D'Esposito, M., Tags: Metabolism, Neuroscience Articles Source Type: research

Non-cell-autonomous function of DR6 in Schwann cell proliferation
Death receptor 6 (DR6) is an orphan member of the TNF receptor superfamily and controls cell death and differentiation in a cell-autonomous manner in different cell types. Here, we report an additional non-cell-autonomous function for DR6 in the peripheral nervous system (PNS). DR6-knockout (DR6 KO) mice showed precocious myelination in the PNS. Using an in vitro myelination assay, we demonstrate that neuronal DR6 acts in trans on Schwann cells (SCs) and reduces SC proliferation and myelination independently of its cytoplasmic death domain. Mechanistically, DR6 was found to be cleaved in neurons by "a disintegrin...
Source: EMBO Journal - April 3, 2018 Category: Molecular Biology Authors: Colombo, A., Hsia, H.-E., Wang, M., Kuhn, P.-H., Brill, M. S., Canevazzi, P., Feederle, R., Taveggia, C., Misgeld, T., Lichtenthaler, S. F. Tags: Neuroscience Articles Source Type: research

The pseudophosphatase STYX targets the F-box of FBXW7 and inhibits SCFFBXW7 function
(Source: EMBO Journal)
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Reiterer, V., Figueras-Puig, C., Le Guerroue, F., Confalonieri, S., Vecchi, M., Jalapothu, D., Kanse, S. M., Deshaies, R. J., Di Fiore, P. P., Behrends, C., Farhan, H. Tags: Corrigendum Source Type: research

No way out: when RNA elements promote nuclear retention
The proper localization of RNA transcripts is a highly controlled and fine-tuned process. Indeed, regulation of RNA trafficking is mediated by both cis-acting elements and trans-acting factors, and defects in either mechanism have been associated with disease. Identifying the RNA sequence motifs that determine cellular localization for a given transcript therefore represents an important and challenging task. A new study from Shukla et al in The EMBO Journal—along with related work from Lubelsky and Ulitsky published elsewhere—describes a new screen that uses hybrid RNAs with barcoded oligonucleotides to i...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Agostini, F., Ule, J., Zagalak, J. A. Tags: Methods & Resources, RNA Biology, Systems & Computational Biology News [amp ] Views Source Type: research

High-throughput identification of RNA nuclear enrichment sequences
We examined a pool of 11,969 oligos densely tiling 38 human lncRNAs that were fused to a cytosolic transcript. After cell fractionation and barcode sequencing, we identified 109 unique RNA regions that significantly enriched this cytosolic transcript in the nucleus including a cytosine-rich motif. These nuclear enrichment sequences are highly conserved and over-represented in global nuclear fractionation sequencing. Importantly, many of these regions were independently validated by single-molecule RNA fluorescence in situ hybridization. Overall, we demonstrate the utility of MPRNA for future investigation of RNA-based...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Shukla, C. J., McCorkindale, A. L., Gerhardinger, C., Korthauer, K. D., Cabili, M. N., Shechner, D. M., Irizarry, R. A., Maass, P. G., Rinn, J. L. Tags: Methods & Resources, RNA Biology, Systems & Computational Biology Source Type: research

LPS targets host guanylate-binding proteins to the bacterial outer membrane for non-canonical inflammasome activation
Pathogenic and commensal Gram-negative bacteria produce and release outer membrane vesicles (OMVs), which present several surface antigens and play an important role for bacterial pathogenesis. OMVs also modulate the host immune system, which makes them attractive as vaccine candidates. At the cellular level, OMVs are internalized by macrophages and deliver lipopolysaccharide (LPS) into the host cytosol, thus activating the caspase-11 non-canonical inflammasome. Here, we show that OMV-induced inflammasome activation requires TLR4-TRIF signaling, the production of type I interferons, and the action of guanylate-binding prot...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Santos, J. C., Dick, M. S., Lagrange, B., Degrandi, D., Pfeffer, K., Yamamoto, M., Meunier, E., Pelczar, P., Henry, T., Broz, P. Tags: Immunology Articles Source Type: research

Molecular basis for sterol transport by StART-like lipid transfer domains
Lipid transport proteins at membrane contact sites, where two organelles are closely apposed, play key roles in trafficking lipids between cellular compartments while distinct membrane compositions for each organelle are maintained. Understanding the mechanisms underlying non-vesicular lipid trafficking requires characterization of the lipid transporters residing at contact sites. Here, we show that the mammalian proteins in the lipid transfer proteins anchored at a membrane contact site (LAM) family, called GRAMD1a-c, transfer sterols with similar efficiency as the yeast orthologues, which have known roles in sterol trans...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Horenkamp, F. A., Valverde, D. P., Nunnari, J., Reinisch, K. M. Tags: Membrane & Intracellular Transport, Structural Biology Articles Source Type: research

A single nucleotide incorporation step limits human telomerase repeat addition activity
We report here that the unique template-embedded pause signal restricts the first nucleotide incorporation for each repeat synthesized, imparting a significantly greater KM. This slow nucleotide incorporation step drastically limits repeat addition processivity and rate under physiological conditions, which is alleviated with augmented concentrations of dGTP or dGDP, and not with dGMP nor other nucleotides. The activity stimulation by dGDP is due to nucleoside diphosphates functioning as substrates for telomerase. Converting the first nucleotide of the repeat synthesized from dG to dA through the telomerase template mutati...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Chen, Y., Podlevsky, J. D., Logeswaran, D., Chen, J. J.-L. Tags: DNA Replication, Repair & Recombination Articles Source Type: research

Prion-like protein aggregates exploit the RHO GTPase to cofilin-1 signaling pathway to enter cells
Protein aggregation is a hallmark of diverse neurodegenerative diseases. Multiple lines of evidence have revealed that protein aggregates can penetrate inside cells and spread like prions. How such aggregates enter cells remains elusive. Through a focused siRNA screen targeting genes involved in membrane trafficking, we discovered that mutant SOD1 aggregates, like viruses, exploit cofilin-1 to remodel cortical actin and enter cells. Upstream of cofilin-1, signalling from the RHO GTPase and the ROCK1 and LIMK1 kinases controls cofilin-1 activity to remodel actin and modulate aggregate entry. In the spinal cord of symptomati...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Zhong, Z., Grasso, L., Sibilla, C., Stevens, T. J., Barry, N., Bertolotti, A. Tags: Cell Adhesion, Polarity & Cytoskeleton, Membrane & Intracellular Transport, Protein Biosynthesis & Quality Control Articles Source Type: research

HP1{alpha} targets the chromosomal passenger complex for activation at heterochromatin before mitotic entry
The chromosomal passenger complex (CPC) is directed to centromeres during mitosis via binding to H3T3ph and Sgo1. Whether and how heterochromatin protein 1α (HP1α) influences CPC localisation and function during mitotic entry is less clear. Here, we alter HP1α dynamics by fusing it to a CENP-B DNA-binding domain. Tethered HP1 strongly recruits the CPC, destabilising kinetochore–microtubule interactions and activating the spindle assembly checkpoint. During mitotic exit, the tethered HP1 traps active CPC at centromeres. These HP1-CPC clusters remain catalytically active throughout the subsequent cell...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Ruppert, J. G., Samejima, K., Platani, M., Molina, O., Kimura, H., Jeyaprakash, A. A., Ohta, S., Earnshaw, W. C. Tags: Cell Cycle, Chromatin, Epigenetics, Genomics & Functional Genomics Articles Source Type: research

Structural basis of siRNA recognition by TRBP double-stranded RNA binding domains
The accurate cleavage of pre-micro(mi)RNAs by Dicer and mi/siRNA guide strand selection are important steps in forming the RNA-induced silencing complex (RISC). The role of Dicer binding partner TRBP in these processes remains poorly understood. Here, we solved the solution structure of the two N-terminal dsRNA binding domains (dsRBDs) of TRBP in complex with a functionally asymmetric siRNA using NMR, EPR, and single-molecule spectroscopy. We find that siRNA recognition by the dsRBDs is not sequence-specific but rather depends on the RNA shape. The two dsRBDs can swap their binding sites, giving rise to two equally populat...
Source: EMBO Journal - March 15, 2018 Category: Molecular Biology Authors: Masliah, G., Maris, C., König, S. L., Yulikov, M., Aeschimann, F., Malinowska, A. L., Mabille, J., Weiler, J., Holla, A., Hunziker, J., Meisner-Kober, N., Schuler, B., Jeschke, G., Allain, F. H.-T. Tags: RNA Biology, Structural Biology Articles Source Type: research

STIM1 (c)AMPs up melanogenesis
STIM1 and STIM2 are endoplasmic reticulum Ca2+ sensors that serve multi-faceted roles in signal transduction in a wide variety of different cell types. In this issue of The EMBO Journal, Motiani et al define and characterize the ability of STIM1 to control cAMP generation in a new context, melanin production (Motiani et al, 2018), offering new insights into the physiological role of STIM1. (Source: EMBO Journal)
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Soboloff, J., Gligorijevic, B., Zaidi, M. R. Tags: Development & Differentiation, Physiology, Signal Transduction News [amp ] Views Source Type: research

p62 filaments capture and present ubiquitinated cargos for autophagy
The removal of misfolded, ubiquitinated proteins is an essential part of the protein quality control. The ubiquitin-proteasome system (UPS) and autophagy are two interconnected pathways that mediate the degradation of such proteins. During autophagy, ubiquitinated proteins are clustered in a p62-dependent manner and are subsequently engulfed by autophagosomes. However, the nature of the protein substrates targeted for autophagy is unclear. Here, we developed a reconstituted system using purified components and show that p62 and ubiquitinated proteins spontaneously coalesce into larger clusters. Efficient cluster formation ...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Zaffagnini, G., Savova, A., Danieli, A., Romanov, J., Tremel, S., Ebner, M., Peterbauer, T., Sztacho, M., Trapannone, R., Tarafder, A. K., Sachse, C., Martens, S. Tags: Autophagy & Cell Death, Post-translational Modifications, Proteolysis & Proteomics Articles Source Type: research

Chk1 and 14-3-3 proteins inhibit atypical E2Fs to prevent a permanent cell cycle arrest
The atypical E2Fs, E2F7 and E2F8, act as potent transcriptional repressors of DNA replication genes providing them with the ability to induce a permanent S-phase arrest and suppress tumorigenesis. Surprisingly in human cancer, transcript levels of atypical E2Fs are frequently elevated in proliferating cancer cells, suggesting that the tumor suppressor functions of atypical E2Fs might be inhibited through unknown post-translational mechanisms. Here, we show that atypical E2Fs can be directly phosphorylated by checkpoint kinase 1 (Chk1) to prevent a permanent cell cycle arrest. We found that 14-3-3 protein isoforms interact ...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Yuan, R., Vos, H. R., van Es, R. M., Chen, J., Burgering, B. M., Westendorp, B., de Bruin, A. Tags: Cell Cycle, DNA Replication, Repair & Recombination Articles Source Type: research

AMPK promotes survival of c-Myc-positive melanoma cells by suppressing oxidative stress
Although c-Myc is essential for melanocyte development, its role in cutaneous melanoma, the most aggressive skin cancer, is only partly understood. Here we used the NrasQ61KINK4a–/– mouse melanoma model to show that c-Myc is essential for tumor initiation, maintenance, and metastasis. c-Myc-expressing melanoma cells were preferentially found at metastatic sites, correlated with increased tumor aggressiveness and high tumor initiation potential. Abrogation of c-Myc caused apoptosis in primary murine and human melanoma cells. Mechanistically, c-Myc-positive melanoma cells activated and became dependent on the met...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Kfoury, A., Armaro, M., Collodet, C., Sordet-Dessimoz, J., Giner, M. P., Christen, S., Moco, S., Leleu, M., de Leval, L., Koch, U., Trumpp, A., Sakamoto, K., Beermann, F., Radtke, F. Tags: Cancer Articles Source Type: research

STIM1 activation of adenylyl cyclase 6 connects Ca2+ and cAMP signaling during melanogenesis
Endoplasmic reticulum (ER)–plasma membrane (PM) junctions form functionally active microdomains that connect intracellular and extracellular environments. While the key role of these interfaces in maintenance of intracellular Ca2+ levels has been uncovered in recent years, the functional significance of ER-PM junctions in non-excitable cells has remained unclear. Here, we show that the ER calcium sensor protein STIM1 (stromal interaction molecule 1) interacts with the plasma membrane-localized adenylyl cyclase 6 (ADCY6) to govern melanogenesis. The physiological stimulus α-melanocyte-stimulating hormone (&alpha...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Motiani, R. K., Tanwar, J., Raja, D. A., Vashisht, A., Khanna, S., Sharma, S., Srivastava, S., Sivasubbu, S., Natarajan, V. T., Gokhale, R. S. Tags: Development & Differentiation, Physiology, Signal Transduction Articles Source Type: research

CD1d-mediated lipid presentation by CD11c+ cells regulates intestinal homeostasis
Intestinal homeostasis relies on a continuous dialogue between the commensal bacteria and the immune system. Natural killer T (NKT) cells, which recognize CD1d-restricted microbial lipids and self-lipids, contribute to the regulation of mucosal immunity, yet the mechanisms underlying their functions remain poorly understood. Here, we demonstrate that NKT cells respond to intestinal lipids and CD11c+ cells (including dendritic cells (DCs) and macrophages) are essential to mediate lipid presentation within the gut ultimately controlling intestinal NKT cell homeostasis and activation. Conversely, CD1d and NKT cells participat...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Saez de Guinoa, J., Jimeno, R., Gaya, M., Kipling, D., Garzon, M. J., Dunn-Walters, D., Ubeda, C., Barral, P. Tags: Immunology Articles Source Type: research

A single N-terminal phosphomimic disrupts TDP-43 polymerization, phase separation, and RNA splicing
TDP-43 is an RNA-binding protein active in splicing that concentrates into membraneless ribonucleoprotein granules and forms aggregates in amyotrophic lateral sclerosis (ALS) and Alzheimer's disease. Although best known for its predominantly disordered C-terminal domain which mediates ALS inclusions, TDP-43 has a globular N-terminal domain (NTD). Here, we show that TDP-43 NTD assembles into head-to-tail linear chains and that phosphomimetic substitution at S48 disrupts TDP-43 polymeric assembly, discourages liquid–liquid phase separation (LLPS) in vitro, fluidizes liquid–liquid phase separated nuclear TDP-...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Wang, A., Conicella, A. E., Schmidt, H. B., Martin, E. W., Rhoads, S. N., Reeb, A. N., Nourse, A., Ramirez Montero, D., Ryan, V. H., Rohatgi, R., Shewmaker, F., Naik, M. T., Mittag, T., Ayala, Y. M., Fawzi, N. L. Tags: Neuroscience, Protein Biosynthesis & Quality Control, RNA Biology Articles Source Type: research

Actin retrograde flow controls natural killer cell response by regulating the conformation state of SHP-1
Natural killer (NK) cells are a powerful weapon against viral infections and tumor growth. Although the actin–myosin (actomyosin) cytoskeleton is crucial for a variety of cellular processes, the role of mechanotransduction, the conversion of actomyosin mechanical forces into signaling cascades, was never explored in NK cells. Here, we demonstrate that actomyosin retrograde flow (ARF) controls the immune response of primary human NK cells through a novel interaction between β-actin and the SH2-domain-containing protein tyrosine phosphatase-1 (SHP-1), converting its conformation state, and thereby regulating NK ce...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Matalon, O., Ben-Shmuel, A., Kivelevitz, J., Sabag, B., Fried, S., Joseph, N., Noy, E., Biber, G., Barda-Saad, M. Tags: Cell Adhesion, Polarity & Cytoskeleton, Immunology, Signal Transduction Articles Source Type: research

IRES-mediated translation of cofilin regulates axonal growth cone extension and turning
In neuronal development, dynamic rearrangement of actin promotes axonal growth cone extension, and spatiotemporal translation of local mRNAs in response to guidance cues directs axonal growth cone steering, where cofilin plays a critical role. While regulation of cofilin activity is well studied, regulatory mechanism for cofilin mRNA translation in neurons is unknown. In eukaryotic cells, proteins can be synthesized by cap-dependent or cap-independent mechanism via internal ribosome entry site (IRES)-mediated translation. IRES-mediated translation has been reported in various pathophysiological conditions, but its role in ...
Source: EMBO Journal - March 1, 2018 Category: Molecular Biology Authors: Choi, J.-H., Wang, W., Park, D., Kim, S.-H., Kim, K.-T., Min, K.-T. Tags: Neuroscience, Protein Biosynthesis & Quality Control, RNA Biology Articles Source Type: research

Shuttle mission in the mitochondrial intermembrane space
Lipid trafficking is essential for biogenesis and maintenance of eukaryotic organelles. In this issue of The EMBO Journal, Saita et al (2018) revealed that proteolytic processing by the rhomboid protease PARL in the mitochondrial inner membrane facilitates partitioning of START domain-containing protein STARD7 to the cytosol and mitochondrial intermembrane space. STARD7 in the mitochondrial intermembrane space functions as a lipid transfer protein to shuttle phosphatidylcholine from the outer membrane to the inner membrane. (Source: EMBO Journal)
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Endo, T., Tamura, Y. Tags: Membrane & Intracellular Transport, Metabolism News [amp ] Views Source Type: research

How to navigate counter dogmatic research findings
(Source: EMBO Journal)
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Prochiantz, A. Tags: Development & Differentiation, Transcription Opinion Source Type: research

Sorting out "non-canonical" autophagy
Canonically, LC3 lipidation has been associated with autophagy pathways but it becomes increasingly clear that this modification can also occur during autophagy-unrelated processes. In this issue, Florey and colleagues find that the WD40 domain of ATG16L1 is dispensable for LC3 lipidation during starvation-induced autophagy but required for its lipidation during several other membrane-based processes that are different from autophagy. This finding opens the door for the analysis of the functions of LC3 lipidation in these pathways. (Source: EMBO Journal)
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Fracchiolla, D., Martens, S. Tags: Autophagy & Cell Death, Membrane & Intracellular Transport News [amp ] Views Source Type: research

PARL partitions the lipid transfer protein STARD7 between the cytosol and mitochondria
Intramembrane-cleaving peptidases of the rhomboid family regulate diverse cellular processes that are critical for development and cell survival. The function of the rhomboid protease PARL in the mitochondrial inner membrane has been linked to mitophagy and apoptosis, but other regulatory functions are likely to exist. Here, we identify the START domain-containing protein STARD7 as an intramitochondrial lipid transfer protein for phosphatidylcholine. We demonstrate that PARL-mediated cleavage during mitochondrial import partitions STARD7 to the cytosol and the mitochondrial intermembrane space. Negatively charged amino aci...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Saita, S., Tatsuta, T., Lampe, P. A., König, T., Ohba, Y., Langer, T. Tags: Membrane & Intracellular Transport Articles Source Type: research

The WD40 domain of ATG16L1 is required for its non-canonical role in lipidation of LC3 at single membranes
A hallmark of macroautophagy is the covalent lipidation of LC3 and insertion into the double-membrane phagophore, which is driven by the ATG16L1/ATG5-ATG12 complex. In contrast, non-canonical autophagy is a pathway through which LC3 is lipidated and inserted into single membranes, particularly endolysosomal vacuoles during cell engulfment events such as LC3-associated phagocytosis. Factors controlling the targeting of ATG16L1 to phagophores are dispensable for non-canonical autophagy, for which the mechanism of ATG16L1 recruitment is unknown. Here we show that the WD repeat-containing C-terminal domain (WD40 CTD) of ATG16L...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Fletcher, K., Ulferts, R., Jacquin, E., Veith, T., Gammoh, N., Arasteh, J. M., Mayer, U., Carding, S. R., Wileman, T., Beale, R., Florey, O. Tags: Autophagy & Cell Death, Immunology Articles Source Type: research

High-resolution transcription maps reveal the widespread impact of roadblock termination in yeast
Transcription termination delimits transcription units but also plays important roles in limiting pervasive transcription. We have previously shown that transcription termination occurs when elongating RNA polymerase II (RNAPII) collides with the DNA-bound general transcription factor Reb1. We demonstrate here that many different DNA-binding proteins can induce termination by a similar roadblock (RB) mechanism. We generated high-resolution transcription maps by the direct detection of RNAPII upon nuclear depletion of two essential RB factors or when the canonical termination pathways for coding and non-coding RNAs are defe...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Candelli, T., Challal, D., Briand, J.-B., Boulay, J., Porrua, O., Colin, J., Libri, D. Tags: RNA Biology, Transcription Articles Source Type: research

The RIG-I-like receptor LGP2 inhibits Dicer-dependent processing of long double-stranded RNA and blocks RNA interference in mammalian cells
In vertebrates, the presence of viral RNA in the cytosol is sensed by members of the RIG-I-like receptor (RLR) family, which signal to induce production of type I interferons (IFN). These key antiviral cytokines act in a paracrine and autocrine manner to induce hundreds of interferon-stimulated genes (ISGs), whose protein products restrict viral entry, replication and budding. ISGs include the RLRs themselves: RIG-I, MDA5 and, the least-studied family member, LGP2. In contrast, the IFN system is absent in plants and invertebrates, which defend themselves from viral intruders using RNA interference (RNAi). In RNAi, the endo...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: van der Veen, A. G., Maillard, P. V., Schmidt, J. M., Lee, S. A., Deddouche-Grass, S., Borg, A., Kjaer, S., Snijders, A. P., Reis e Sousa, C. Tags: Immunology, RNA Biology Articles Source Type: research

PlexinD1 signaling controls morphological changes and migration termination in newborn neurons
Newborn neurons maintain a very simple, bipolar shape, while they migrate from their birthplace toward their destinations in the brain, where they differentiate into mature neurons with complex dendritic morphologies. Here, we report a mechanism by which the termination of neuronal migration is maintained in the postnatal olfactory bulb (OB). During neuronal deceleration in the OB, newborn neurons transiently extend a protrusion from the proximal part of their leading process in the resting phase, which we refer to as a filopodium-like lateral protrusion (FLP). The FLP formation is induced by PlexinD1 downregulation and lo...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Sawada, M., Ohno, N., Kawaguchi, M., Huang, S.-h., Hikita, T., Sakurai, Y., Bang Nguyen, H., Quynh Thai, T., Ishido, Y., Yoshida, Y., Nakagawa, H., Uemura, A., Sawamoto, K. Tags: Cell Adhesion, Polarity & Cytoskeleton, Neuroscience Articles Source Type: research

ERAD-dependent control of the Wnt secretory factor Evi
Active regulation of protein abundance is an essential strategy to modulate cellular signaling pathways. Within the Wnt signaling cascade, regulated degradation of β-catenin by the ubiquitin-proteasome system (UPS) affects the outcome of canonical Wnt signaling. Here, we found that abundance of the Wnt cargo receptor Evi (Wls/GPR177), which is required for Wnt protein secretion, is also regulated by the UPS through endoplasmic reticulum (ER)-associated degradation (ERAD). In the absence of Wnt ligands, Evi is ubiquitinated and targeted for ERAD in a VCP-dependent manner. Ubiquitination of Evi involves the E2-conjugati...
Source: EMBO Journal - February 15, 2018 Category: Molecular Biology Authors: Glaeser, K., Urban, M., Fenech, E., Voloshanenko, O., Kranz, D., Lari, F., Christianson, J. C., Boutros, M. Tags: Membrane & Intracellular Transport, Protein Biosynthesis & Quality Control, Signal Transduction Articles Source Type: research