57 research outputs found
First person – Varun Jayeshkumar Shah
No full textABSTRACT
First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping early-career researchers promote themselves alongside their papers. Varun Jayeshkumar Shah is the first author on ‘CRL7SMU1 E3 ligase complex-driven H2B ubiquitination functions in sister chromatid cohesion by regulating SMC1 expression’, published in Journal of Cell Science. Varun is a PhD student in the lab of Dr Subbareddy Maddika at the Centre for DNA Fingerprinting and Diagnostics (CDFD), Hyderabad, India, investigating the role of LisH-domain-containing proteins in the assembly of multi-subunit E3 ligase complexes.</jats:p
Interplay between the phosphatase PHLPP1 and E3 ligase RNF41 stimulates proper kinetochore assembly via the outer-kinetochore protein SGT1
No full textKinetochores link chromosomes to spindle microtubules and are essential for accurate chromosome segregation during cell division. Kinetochores assemble at the centromeric region of chromosomes as a multiprotein complex. However, the molecular mechanisms of kinetochore assembly have not yet been fully elucidated. In this study, we identified pleckstrin homology domain and leucine-rich repeat protein phosphatase 1 (PHLPP1) as a regulatory phosphatase that facilitates proper kinetochore assembly. We found that PHLPP1 interacted with the essential outer-kinetochore protein SGT1 and stabilized its protein levels. Loss of PHLPP1 from cells led to SGT1 degradation and thereby caused defective kinetochore assembly. We also found that the ring finger protein 41 (RNF41) as an E3 ligase ubiquitinated and degraded SGT1 in a phosphorylation-dependent manner. PHLPP1 dephosphorylated SGT1 at four conserved residues (Ser-17, Ser-249, Ser-289, and Thr-233) and thereby prevented SGT1 from associating with RNF41, in turn, countering SGT1 degradation. Importantly, depletion of RNF41 or expression of a non-phosphorylatable SGT1 mutant rescued the kinetochore defects caused by the loss of PHLPP1. Taken together, our results suggest that PHLPP1 plays an important role in the assembly of kinetochores by counteracting RNF41-mediated SGT1 degradation
Protein kinase DYRK2 is a scaffold that facilitates assembly of an E3 ligase
No full textProtein kinases have central functions in various cellular signal transduction pathways through their substrate phosphorylation. Here we show that a protein kinase, DYRK2, has unexpected role as a scaffold for an E3 ubiquitin ligase complex. DYRK2 associates with an E3 ligase complex containing EDD, DDB1 and VPRBP proteins (EDVP complex). Strikingly, DYRK2 serves as a scaffold for the EDVP complex, because small-interfering-RNA-mediated depletion of DYRK2 disrupts the formation of the EDD-DDB1-VPRBP complex. Although the kinase activity of DYRK2 is dispensable for its ability to mediate EDVP complex formation, it is required for the phosphorylation and subsequent degradation of its downstream substrate, katanin p60. Collectively, our results reveal a new type of E3-ubiquitin ligase complex in humans that depends on a protein kinase for complex formation as well as for the subsequent phosphorylation, ubiquitylation and degradation of their substrates
PTEN Regulates Glucose Transporter Recycling by Impairing SNX27 Retromer Assembly
No full textSummary: The tumor suppressor PTEN executes cellular functions predominantly through its phosphatase activity. Here we identified a phosphatase-independent role for PTEN during vesicular trafficking of the glucose transporter GLUT1. PTEN physically interacts with SNX27, a component of the retromer complex that recycles transmembrane receptors such as GLUT1 from endosomes to the plasma membrane. PTEN binding with SNX27 prevents GLUT1 accumulation at the plasma membrane because of defective recycling and thus reduces cellular glucose uptake. Mechanistically, PTEN blocks the association of SNX27 with VPS26 and thereby hinders assembly of a functional retromer complex during the receptor recycling process. Importantly, we found a PTEN somatic mutation (T401I) that is defective in disrupting the association between SNX27 and VPS26, suggesting a critical role for PTEN in controlling optimal GLUT1 levels at the membrane to prevent tumor progression. Together, our results reveal a fundamental role of PTEN in the regulation of the SNX27 retromer pathway, which governs glucose transport and might contribute to PTEN tumor suppressor function. : Shinde et al. identify a critical role for the tumor suppressor PTEN in the regulation of glucose uptake by cells. PTEN binds SNX27 and hinders its access to the VPS26 retromer complex, preventing recycling of the glucose transporter GLUT1 to the plasma membrane, which leads to impaired cellular glucose uptake. Keywords: PTEN, GLUT1, retromer, SNX27, endosomal recyclin
Cellular Dynamics Controlled by Phosphatases
No full textProtein phosphorylation, a fundamental post-translation modification that acts as a backbone of signaling networks, is essential for multiple aspects of eukaryote physiology. Phosphorylation status of a substrate is dependent on opposing activities of two distinct enzymes, where the relevant kinase catalyzes the modification and is reversed by a phosphatase. Historically, kinases have been at the research forefront; however, phosphatases have gained importance with many studies revealing predominant roles for these enzymes in controlling the cellular responses. Phosphatases are known to attenuate or amplify signaling by operating both as early, as well as delayed regulators of signal transduction. This review is focused on describing the versatile roles of phosphatases in controlling different cellular pathways through their spatio-temporal dynamics during signaling
Post translational modifications of Rab GTPases
No full textRab GTPases, the highly conserved members of Ras GTPase superfamily are central players in the vesicular trafficking. They are critically involved in intracellular trafficking pathway, beginning from formation of vesicles on donor membranes, defining trafficking specificity to facilitating vesicle docking on target membranes. Given the dynamic roles of Rabs during different stages of vesicular trafficking, mechanisms for their spatial and temporal regulation are crucial for normal cellular function. Regulation of Rab GTPase activity, localization and function has always been focused in and around the association of GDP dissociation inhibitor (GDI), Guanine nucleotide Exchange Factor (GEFs) and GTPase accelerating protein (GAP) to Rabs. However, several recent studies have highlighted the importance of different post-translational modifications in regulation of Rab activation and function. This review provides a summary of various post translational modifications (PTMs) and their significance to regulate localization and function of different Rabs
A modification switch on a molecular switch: Phosphoregulation of Rab7 during endosome maturation
No full textRab GTPases, the highly conserved members of Ras GTPase superfamily are the pivotal regulators of vesicle-mediated trafficking. Rab GTPases, each with a specific subcellular localization, exert tremendous control over various aspects of vesicular transport, identity and dynamics. Several lines of research have established that GDI, GEFs and GAPs are the critical players to orchestrate Rab GTPase activity and function. The importance of post translational modifications in Rab GTPase functional regulation is poorly or not yet been addressed except for prenylation. Our recent study has revealed a novel dephosphorylation dependent regulatory mechanism for Rab7 activity and function. We have shown the importance of PTEN mediated dephosphorylation of Rab7 on highly conserved S72 and Y183 residues, which is essential for its GDI mediated membrane targeting and further activation by GEF. In conclusion, our study highlighted the importance of a phosphorylation/dephosphorylation switch in controlling timely Rab7 localization and activity on endosomes
HACE1 mediated K27 ubiquitin linkage leads to YB-1 protein secretion
No full textUbiquitination is an important post-translational modification that is implicated in controlling almost every biological process by targeting cellular proteins to degradation. While the importance of ubiquitination in controlling the fate and the intracellular functions of various proteins was widely studied, its role in extracellular protein secretion has been unexplored so far. In this study, by using YB-1 (Y-box Binding protein 1) as a model protein, we showed that ubiquitination is required for its extracellular secretion. We also identified HACE1 as a specific E3 ligase that polyubiquitinates YB-1 through non-canonical K27 linked ubiquitin chains. Formation of these ubiquitin linkages on YB-1 is necessary for its interaction with Tumor Susceptibility Gene 101 (TSG101), a component of the Multi-Vesicular Body (MVB) pathway, which facilitates its secretion. Finally, we demonstrated that extracellular secreted YB-1 is a functional protein that acts to inhibit Transforming Growth Factor-Beta mediated epithelial to mesenchymal transition. In summary, we identified a novel functional role for non-canonical ubiquitin linkages in mediating protein secretion
CRL7SMU1 E3 ligase complex driven H2B ubiquitination functions in sister chromatid cohesion by regulating SMC1 expression
No full textCullin–RING E3 ligases (CRLs) control broad range of biological processes by ubiquitinating numerous cellular substrates. However, role of CRL E3 ligases in chromatid cohesion is unknown. In this study, we identified a new CRL type E3 ligase (designated as CRL7SMU1 complex) that has an essential role in maintenance of chromatid cohesion. We demonstrate that SMU1, DDB1, CUL7 and RNF40 as integral components of this complex. SMU1 by acting as a substrate recognition module, binds to H2B and mediates monoubiquitination at K120 site through CRL7SMU1 E3 ligase complex. Depletion of CRL7SMU1 leads to loss of H2B ubiquitination at SMC1a locus and thus subsequently compromised SMC1a expression in cells. Knock down of CRL7SMU1 components or loss of H2B ubiquitination leads to defective sister chromatid cohesion, which is rescued by restoration of SMC1a expression. Together, our results unveil an important role of CRL7SMU1 E3 ligase in promoting H2B ubiquitination for maintenance of sister chromatid cohesion during mitosis.</jats:p
WD Repeat Protein WDR48 in Complex with Deubiquitinase USP12 Suppresses Akt-dependent Cell Survival Signaling by Stabilizing PH Domain Leucine-rich Repeat Protein Phosphatase 1 (PHLPP1)
No full textPHLPP1 (PH domain leucine-rich repeat protein phosphatase 1) is a protein-serine/threonine phosphatase and a negative regulator of the PI3-kinase/Akt pathway. Although its function as a suppressor of tumor cell growth has been established, the mechanism of its regulation is not completely understood. In this study, by utilizing the tandem affinity purification approach we have identified WDR48 and USP12 as novel PHLPP1-associated proteins. The WDR48·USP12 complex deubiquitinates PHLPP1 and thereby enhances its protein stability. Similar to PHLPP1 function, WDR48 and USP12 negatively regulate Akt activation and thus promote cellular apoptosis. Functionally, we show that WDR48 and USP12 suppress proliferation of tumor cells. Importantly, we found a WDR48 somatic mutation (L580F) that is defective in stabilizing PHLPP1 in colorectal cancers, supporting a WDR48 role in tumor suppression. Together, our results reveal WDR48 and USP12 as novel PHLPP1 regulators and potential suppressors of tumor cell survival
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