Dual Regulation of the Mitotic Exit Network (MEN) by PP2A-Cdc55 Phosphatase

Baró, Bàrbara; Rodriguez-Rodriguez, José-Antonio; Calabria, Inés; Hernáez, María Luisa; Gil, Concha; Queralt, Ethel

doi:10.1371/journal.pgen.1003966

Cited by 25 publications

(27 citation statements)

References 73 publications

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“…Thus, whether the FEAR is activated or inhibited influences the impact of the chimeric proteins on mitotic exit. PP2A Cdc55 has been recently shown to antagonize the Cdc5-dependent phosphorylation of Bfa1 [ 71 ], thereby providing a mechanistic explanation to our data. The FEAR-mediated partial release of Cdc14 from the nucleolus might also contribute to MEN activation by counteracting the CDK-dependent inhibitory phosphorylation of MEN components [ 63 ].…”

Section: Discussionsupporting

confidence: 60%

“…Critical regulators of Bub2/Bfa1 asymmetry are the polo kinase Cdc5 and the phosphatase PP2A Cdc55 . Cdc5 phosphorylates and inactivates the Bub2/Bfa1 complex leading to Tem1 activation [ 51 , 70 ], whereas PP2A Cdc55 dephosphorylates Bfa1 [ 71 ]. Phosphomimetic mutations in some Cdc5-dependent Bfa1 phosphorylation sites, as well as loss of PP2A Cdc55 , are sufficient to induce premature asymmetry of Bub2/Bfa1 at SPBs, whereas Cdc5 inactivation or phospho-ablating mutations in Bfa1 lead to its persistent symmetry [ 62 , 71 ].…”

Section: Discussionmentioning

confidence: 99%

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Asymmetry of the Budding Yeast Tem1 GTPase at Spindle Poles Is Required for Spindle Positioning But Not for Mitotic Exit

et al. 2015

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The asymmetrically dividing yeast S. cerevisiae assembles a bipolar spindle well after establishing the future site of cell division (i.e., the bud neck) and the division axis (i.e., the mother-bud axis). A surveillance mechanism called spindle position checkpoint (SPOC) delays mitotic exit and cytokinesis until the spindle is properly positioned relative to the mother-bud axis, thereby ensuring the correct ploidy of the progeny. SPOC relies on the heterodimeric GTPase-activating protein Bub2/Bfa1 that inhibits the small GTPase Tem1, in turn essential for activating the mitotic exit network (MEN) kinase cascade and cytokinesis. The Bub2/Bfa1 GAP and the Tem1 GTPase form a complex at spindle poles that undergoes a remarkable asymmetry during mitosis when the spindle is properly positioned, with the complex accumulating on the bud-directed old spindle pole. In contrast, the complex remains symmetrically localized on both poles of misaligned spindles. The mechanism driving asymmetry of Bub2/Bfa1/Tem1 in mitosis is unclear. Furthermore, whether asymmetry is involved in timely mitotic exit is controversial. We investigated the mechanism by which the GAP Bub2/Bfa1 controls GTP hydrolysis on Tem1 and generated a series of mutants leading to constitutive Tem1 activation. These mutants are SPOC-defective and invariably lead to symmetrical localization of Bub2/Bfa1/Tem1 at spindle poles, indicating that GTP hydrolysis is essential for asymmetry. Constitutive tethering of Bub2 or Bfa1 to both spindle poles impairs SPOC response but does not impair mitotic exit. Rather, it facilitates mitotic exit of MEN mutants, likely by increasing the residence time of Tem1 at spindle poles where it gets active. Surprisingly, all mutant or chimeric proteins leading to symmetrical localization of Bub2/Bfa1/Tem1 lead to increased symmetry at spindle poles of the Kar9 protein that mediates spindle positioning and cause spindle misalignment. Thus, asymmetry of the Bub2/Bfa1/Tem1 complex is crucial to control Kar9 distribution and spindle positioning during mitosis.

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Section: Discussionsupporting

confidence: 60%

Section: Discussionmentioning

confidence: 99%

Asymmetry of the Budding Yeast Tem1 GTPase at Spindle Poles Is Required for Spindle Positioning But Not for Mitotic Exit

et al. 2015

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“…Thus, Zds1/2 are common PP2A Cdc55 modulators, participating in both entry and exit from mitosis. It has recently been described that PP2A Cdc55 downregulation in anaphase also initiates the Mitotic Exit Network (MEN) by dephosphorylating the MEN components Bfa1 and Mob1 [ 43 ]. In addition, PP2A Cdc55 downregulation at anaphase-onset facilitates separase proteolytic activity towards Scc1, which triggers sister-chromatid segregation [ 44 ].…”

Section: Introductionmentioning

confidence: 99%

SILAC-based phosphoproteomics reveals new PP2A-Cdc55-regulated processes in budding yeast

et al. 2018

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BackgroundProtein phosphatase 2A (PP2A) is a family of conserved serine/threonine phosphatases involved in several essential aspects of cell growth and proliferation. PP2ACdc55 phosphatase has been extensively related to cell cycle events in budding yeast; however, few PP2ACdc55 substrates have been identified. Here, we performed a quantitative mass spectrometry approach to reveal new substrates of PP2ACdc55 phosphatase and new PP2A-related processes in mitotic arrested cells.ResultsWe identified 62 statistically significant PP2ACdc55 substrates involved mainly in actin-cytoskeleton organization. In addition, we validated new PP2ACdc55 substrates such as Slk19 and Lte1, involved in early and late anaphase pathways, and Zeo1, a component of the cell wall integrity pathway. Finally, we constructed docking models of Cdc55 and its substrate Mob1. We found that the predominant interface on Cdc55 is mediated by a protruding loop consisting of residues 84–90, thus highlighting the relevance of these aminoacids for substrate interaction.ConclusionsWe used phosphoproteomics of Cdc55-deficient cells to uncover new PP2ACdc55 substrates and functions in mitosis. As expected, several hyperphosphorylated proteins corresponded to Cdk1-dependent substrates, although other kinases’ consensus motifs were also enriched in our dataset, suggesting that PP2ACdc55 counteracts and regulates other kinases distinct from Cdk1. Indeed, Pkc1 emerged as a novel node of PP2ACdc55 regulation, highlighting a major role of PP2ACdc55 in actin cytoskeleton and cytokinesis, gene ontology terms significantly enriched in the PP2ACdc55-dependent phosphoproteome.

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“…4 In budding yeast, cells lacking Cdc55 (a regulatory B subunit) exhibit multiple cell cycle defects, including mitotic entry, [5][6][7][8][9][10] spindle assembly checkpoint (SAC), 5,[11][12][13] and precocious mitotic exit. 12,[14][15][16][17][18] We have proposed previously that PP2A-Cdc55 activity is largely controlled by its subcellular distribution. 13,16 Cdc55 binding protein Zds1 (zillion different screens) and its homolog Zds2 are cytoplasmic proteins and sequester Cdc55 from the nucleus during mitosis.…”

Section: Introductionmentioning

confidence: 99%