scholarly journals Spatiotemporal coordination of Greatwall-Endos-PP2A promotes mitotic progression

2021 ◽  
Vol 220 (6) ◽  
Author(s):  
Myreille Larouche ◽  
David Kachaner ◽  
Peng Wang ◽  
Karine Normandin ◽  
Damien Garrido ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Regulatory Subunit ◽  
Mitotic Progression ◽  
Mitotic Kinases ◽  
Nuclear Envelope Breakdown ◽  
Spatiotemporal Regulation ◽  
Phosphatase 2A ◽  
Localization Signal ◽  
Mitotic Regulation ◽  
Spatiotemporal Coordination

Mitotic entry involves inhibition of protein phosphatase 2A bound to its B55/Tws regulatory subunit (PP2A-B55/Tws), which dephosphorylates substrates of mitotic kinases. This inhibition is induced when Greatwall phosphorylates Endos, turning it into an inhibitor of PP2A-Tws. How this mechanism operates spatiotemporally in the cell is incompletely understood. We previously reported that the nuclear export of Greatwall in prophase promotes mitotic progression. Here, we examine the importance of the localized activities of PP2A-Tws and Endos for mitotic regulation. We find that Tws shuttles through the nucleus via a conserved nuclear localization signal (NLS), but expression of Tws in the cytoplasm and not in the nucleus rescues the development of tws mutants. Moreover, we show that Endos must be in the cytoplasm before nuclear envelope breakdown (NEBD) to be efficiently phosphorylated by Greatwall and to bind and inhibit PP2A-Tws. Disrupting the cytoplasmic function of Endos before NEBD results in subsequent mitotic defects. Evidence suggests that this spatiotemporal regulation is conserved in humans.

scholarly journals The Ndc80 complex targets Bod1 to human mitotic kinetochores

Open Biology ◽  
2017 ◽  
Vol 7 (11) ◽  
pp. 170099 ◽  
Author(s):  
Katharina Schleicher ◽  
Michael Porter ◽  
Sara ten Have ◽  
Ramasubramanian Sundaramoorthy ◽  
Iain M. Porter ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Regulatory Subunit ◽  
Mitotic Progression ◽  
Mitotic Chromosomes ◽  
Temporal Regulation ◽  
Nuclear Envelope Breakdown ◽  
Sister Chromatids ◽  
Endogenous Protein ◽  
Phosphatase 2A ◽  
Spatio Temporal

Regulation of protein phosphatase activity by endogenous protein inhibitors is an important mechanism to control protein phosphorylation in cells. We recently identified Biorientation defective 1 (Bod1) as a small protein inhibitor of protein phosphatase 2A containing the B56 regulatory subunit (PP2A-B56). This phosphatase controls the amount of phosphorylation of several kinetochore proteins and thus the establishment of load-bearing chromosome-spindle attachments in time for accurate separation of sister chromatids in mitosis. Like PP2A-B56, Bod1 directly localizes to mitotic kinetochores and is required for correct segregation of mitotic chromosomes. In this report, we have probed the spatio-temporal regulation of Bod1 during mitotic progression. Kinetochore localization of Bod1 increases from nuclear envelope breakdown until metaphase. Phosphorylation of Bod1 at threonine 95 (T95), which increases Bod1's binding to and inhibition of PP2A-B56, peaks in prometaphase when PP2A-B56 localization to kinetochores is highest. We demonstrate here that kinetochore targeting of Bod1 depends on the outer kinetochore protein Ndc80 and not PP2A-B56. Crucially, Bod1 depletion functionally affects Ndc80 phosphorylation at the N-terminal serine 55 (S55), as well as a number of other phosphorylation sites within the outer kinetochore, including Knl1 at serine 24 and 60 (S24, S60), and threonine T943 and T1155 (T943, T1155). Therefore, Ndc80 recruits a phosphatase inhibitor to kinetochores which directly feeds forward to regulate Ndc80, and Knl1 phosphorylation, including sites that mediate the attachment of microtubules to kinetochores.

scholarly journals PP2A Phosphatase Activity Is Required for Stress and Tor Kinase Regulation of Yeast Stress Response Factor Msn2p

2004 ◽  
Vol 3 (5) ◽  
pp. 1261-1271 ◽  
Author(s):  
Arti Santhanam ◽  
Alan Hartley ◽  
Katrin Düvel ◽  
James R. Broach ◽  
Stephen Garrett
Keyword(s):  
Protein Function ◽  
Nuclear Export ◽  
Kinase Inhibitor ◽  
Osmotic Shock ◽  
Nuclear Accumulation ◽  
Regulatory Subunit ◽  
Interacting Protein ◽  
Broad Array ◽  
Tor Kinase ◽  
Phosphatase 2A

ABSTRACT In response to stress and nutrient starvation, the Saccharomyces cerevisiae transcription factor Msn2p accumulates in the nucleus and activates expression of a broad array of genes. Here, we analyze the role of the Tor (target of rapamycin) signaling pathway in mediating these responses. Inactivation of the Tor pathway component Tap42p using tap42(Ts) alleles causes a sustained nuclear localization similar to that after the addition of the Tor kinase inhibitor rapamycin. Effects of Tap42p inactivation and rapamycin addition could be suppressed by deletion of TIP41, which encodes a Tap42p-interacting protein. These results support the notion that rapamycin affects Msn2p by inactivating Tap42p function. Tap42p interacts with the catalytic subunit of PP2A (protein phosphatase 2A) and PP2A-like phosphatases. Deletion of either the catalytic or regulatory subunit that forms the PP2A phosphatase complex prevents nuclear accumulation of Msn2p in the tap42(Ts) strain and in wild-type strains treated with rapamycin. These results suggest that Tap42p is an inhibitor of PP2A phosphatase, which in turn inhibits nuclear export of Msn2p. Interestingly, PP2A function is also required for nuclear accumulation of Msn2p in response to stresses, such as heat and osmotic shock, as well as nitrogen (but not glucose) starvation. Thus, PP2A and the Tor kinase pathway transduce stress and nitrogen starvation signals to Msn2p. Finally, Msn2p localization is unaffected by conditional loss of 14-3-3 protein function, ruling out the possibility that 14-3-3 proteins act as a scaffold to sequester Msn2p in the cytoplasm.

scholarly journals AKT regulates mitotic progression of mammalian cells by phosphorylating MASTL

2018 ◽  
Author(s):  
Irfana Reshi ◽  
Misbah Un Nisa ◽  
Umer Farooq ◽  
Syed Qaaifah Gillani ◽  
Sameer Ahmad Bhat ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Mammalian Cells ◽  
Critical Role ◽  
Mitotic Progression ◽  
Oncogenic Potential ◽  
Mitotic Kinases ◽  
Colorectal Cell ◽  
Phosphatase 2A ◽  
Important Regulator ◽  
Regulatory Loop

AbstractMicrotubule associated serine threonine like kinase (MASTL) has been recently identified as an important regulator of mitosis. By inhibiting protein phosphatase 2A, it plays a crucial role in activating one of the most important mitotic kinases known as cyclin dependent kinase1 (CDK1). MASTL has been seen to be up regulated in various types of cancers and is involved in tumor recurrence. It is activated by CDK1 through its auto regulatory loop but the complete mechanism of its activation is still unclear. In this study, we evaluated the regulation of MASTL via AKT during mitosis. Here we report that AKT phosphorylates MASTL at T299 which plays a critical role in its activation. Our results suggest that AKT increases CDK1 mediated phosphorylation and hence activity of MASTL which in turn promotes cell proliferation.. We also show that the oncogenic potential of AKT is augmented by MASTL activation as the AKT mediated oncogenesis in colorectal cell lines can be attenuated by inhibiting and/or silencing MASTL. In brief, we report that AKT has an important role in the progression of mitosis in mammalian cells and it does so through the phosphorylation and activation of MASTL.

scholarly journals AKT Regulates Mitotic Progression of Mammalian Cells by Phosphorylating MASTL, Leading to Protein Phosphatase 2A Inactivation

2020 ◽  
Vol 40 (10) ◽  
Author(s):  
Irfana Reshi ◽  
Misbah Un Nisa ◽  
Umer Farooq ◽  
Syed Qaaifah Gillani ◽  
Sameer Ahmed Bhat ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Mammalian Cells ◽  
Protein Phosphatase 2A ◽  
Critical Role ◽  
Mitotic Progression ◽  
Serine Threonine Kinase ◽  
Threonine Kinase ◽  
Mitotic Kinases ◽  
Colorectal Cell ◽  
Phosphatase 2A

ABSTRACT Microtubule-associated serine/threonine kinase like (MASTL), also known as Greatwall (Gwl) kinase, has an important role in the regulation of mitosis. By inhibiting protein phosphatase 2A (PP2A), it plays a crucial role in activating one of the most important mitotic kinases, known as cyclin-dependent kinase 1 (CDK1). MASTL has been seen to be upregulated in various types of cancers and is also involved in tumor recurrence. It is activated by CDK1 through phosphorylations in the activation/T-loop, but the complete mechanism of its activation is still unclear. Here, we report that AKT phosphorylates MASTL at residue T299, which plays a critical role in its activation. Our results suggest that AKT increases CDK1-mediated phosphorylation and hence the activity of MASTL, which, in turn, promotes mitotic progression through PP2A inhibition. We also show that the oncogenic potential of AKT is augmented by MASTL activation, since AKT-mediated proliferation in colorectal cell lines can be attenuated by inhibiting and/or silencing MASTL. In brief, we report that AKT plays an important role in the progression of mitosis in mammalian cells and that it does so through the phosphorylation and activation of MASTL.

scholarly journals SUMO-1 Modification and Its Role in Targeting the Ran GTPase-activating Protein, RanGAP1, to the Nuclear Pore Complex

1998 ◽  
Vol 140 (3) ◽  
pp. 499-509 ◽  
Author(s):  
Michael J. Matunis ◽  
Jian Wu ◽  
Günter Blobel
Keyword(s):  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Gtpase Activating Protein ◽  
Terminal Domain ◽  
Localization Signal

RanGAP1 is the GTPase-activating protein for Ran, a small ras-like GTPase involved in regulating nucleocytoplasmic transport. In vertebrates, RanGAP1 is present in two forms: one that is cytoplasmic, and another that is concentrated at the cytoplasmic fibers of nuclear pore complexes (NPCs). The NPC-associated form of RanGAP1 is covalently modified by the small ubiquitin-like protein, SUMO-1, and we have recently proposed that SUMO-1 modification functions to target RanGAP1 to the NPC. Here, we identify the domain of RanGAP1 that specifies SUMO-1 modification and demonstrate that mutations in this domain that inhibit modification also inhibit targeting to the NPC. Targeting of a heterologous protein to the NPC depended on determinants specifying SUMO-1 modification and also on additional determinants in the COOH-terminal domain of RanGAP1. SUMO-1 modification and these additional determinants were found to specify interaction between the COOH-terminal domain of RanGAP1 and a region of the nucleoporin, Nup358, between Ran-binding domains three and four. Together, these findings indicate that SUMO-1 modification targets RanGAP1 to the NPC by exposing, or creating, a Nup358 binding site in the COOH-terminal domain of RanGAP1. Surprisingly, the COOH-terminal domain of RanGAP1 was also found to harbor a nuclear localization signal. This nuclear localization signal, and the presence of nine leucine-rich nuclear export signal motifs, suggests that RanGAP1 may shuttle between the nucleus and the cytoplasm.

scholarly journals Staufen1 is imported into the nucleolus via a bipartite nuclear localization signal and several modulatory determinants

2005 ◽  
Vol 393 (1) ◽  
pp. 245-254 ◽  
Author(s):  
Catherine Martel ◽  
Paolo Macchi ◽  
Luc Furic ◽  
Michael A. Kiebler ◽  
Luc Desgroseillers
Keyword(s):  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Nuclear Export ◽  
Mammalian Cells ◽  
Rna Binding ◽  
Binding Activity ◽  
Nuclear Trafficking ◽  
Binding Domains ◽  
Bipartite Nuclear Localization Signal ◽  
Localization Signal

Mammalian Stau1 (Staufen1), a modular protein composed of several dsRBDs (double-stranded RNA-binding domains), is probably involved in mRNA localization. Although Stau1 is mostly described in association with the rough endoplasmic reticulum and ribosomes in the cytoplasm, recent studies suggest that it may transit through the nucleus/nucleolus. Using a sensitive yeast import assay, we show that Stau1 is actively imported into the nucleus through a newly identified bipartite nuclear localization signal. As in yeast, the bipartite nuclear localization signal is necessary for Stau1 nuclear import in mammalian cells. It is also required for Stau1 nucleolar trafficking. However, Stau1 nuclear transit seems to be regulated by mechanisms that involve cytoplasmic retention and/or facilitated nuclear export. Cytoplasmic retention is mainly achieved through the action of dsRBD3, with dsRBD2 playing a supporting role in this function. Similarly, dsRBD3, but not its RNA-binding activity, is critical for Stau1 nucleolar trafficking. The function of dsRBD3 is strengthened or stabilized by the presence of dsRBD4 but prevented by the interdomain between dsRBD2 and dsRBD3. Altogether, these results suggest that Stau1 nuclear trafficking is a highly regulated process involving several determinants. The presence of Stau1 in the nucleus/nucleolus suggests that it may be involved in ribonucleoprotein formation in the nucleus and/or in other nuclear functions not necessarily related to mRNA transport.

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