CDK1-mediated phosphorylation at H2B serine 6 is required for mitotic chromosome segregation

Seibert, Markus; Krüger, Marcus; Watson, Nikolaus A.; Sen, Onur; Daum, John R.; Slotman, Johan A.; Braun, Thomas; Houtsmuller, Adriaan B.; Gorbsky, Gary J.; Jacob, Ralf; Higgins, Jonathan M.G.; Schmitz, M. Lienhard

doi:10.1083/jcb.201806057

Cited by 24 publications

(21 citation statements)

References 72 publications

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“…Interestingly, In MG132-arrested HeLa Tet-On cells, SET largely colocalized with ACA in 93% of kinetochores. Thus, the localization patterns of SET during mitosis are quite similar to the ones of Sgo1 as well as consistent with the findings in a very recent report ( Seibert et al, 2019 ). We then examined the colocalization of SET and Sgo1.…”

Section: Resultssupporting

confidence: 92%

SET binding to Sgo1 inhibits Sgo1–cohesin interactions and promotes chromosome segregation

Yang

Chen

et al. 2019

Journal of Cell Biology

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At anaphase onset, Sgo1 function of cohesion protection must be disabled to allow timely chromosome segregation, but how this is achieved is not fully understood. Here, we show that SET, a known PP2A inhibitor, directly binds to a domain in Sgo1 in close proximity to the cohesin-binding motif. The Sgo1–cohesin binding can be disrupted by SET in a dose-dependent manner in vitro as well as by SET overexpression in cells, suggesting that SET is also an inhibitor to the Sgo1–cohesin binding. Furthermore, the SET binding–deficient Sgo1 mutant fully supports centromeric cohesion protection but delays chromosome segregation, suggesting that the SET–Sgo1 binding is required for timely chromosome segregation. Moreover, overexpression of SET WT, not the Sgo1 binding–deficient mutant, exacerbates the occurrence of cohesion fatigue in MG132-arrested cells. Conversely, SET depletion delays it. Thus, we propose that a major function of SET during mitosis is to disrupt the Sgo1–cohesin interaction, thereby promoting centromeric cohesion de-protection and timely chromosome segregation at anaphase onset.

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Section: Resultssupporting

confidence: 92%

SET binding to Sgo1 inhibits Sgo1–cohesin interactions and promotes chromosome segregation

Yang

Chen

et al. 2019

Journal of Cell Biology

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“…For KiPIK screening, immunoblotting and immunofluorescence microscopy, rabbit polyclonal antibodies to H3T3ph (B8634) 47 , INCENP (P240, Cell Signaling Technology #2807), INCENP-S446ph (Jan-Michael Peters, IMP, Vienna) 57 , INCENP-TSSph (Michael Lampson, University of Pennsylvania) 82 , BCL9L S915ph (Cell Signaling Technology #13325), Neurogranin S36ph (Merck-Millipore, ABN426) and γ-tubulin (AK-15, Sigma, T3320); rabbit monoclonal antibodies to the S-pT-P motif (D73F6; Cell Signaling Technology #5243), Cyclin B1 (D5C10; Cell Signaling Technology #12231) and Vinculin (E1E9V; Cell Signaling Technology #13901); mouse monoclonal antibodies to H3S28ph (CMA315) 83 and phospho-tyrosine (P-Tyr-100; Cell Signaling Technology #9411); and sheep polyclonal antibodies to Aurora B (SAB.1, Stephen Taylor, University of Manchester) 84 and BCL9L (R&D Systems, AF4967) were used. Antibodies used for siRNA screening were mouse monoclonal anti-H3T3ph (16B2) 85 and rabbit polyclonal anti-H2BS6ph 86 . Secondary antibodies were: donkey anti-sheep IgG-HRP (ThermoFisher, A16041), goat antirabbit IgG-HRP (Cell Signaling Technology #7074), horse anti-mouse IgG-HRP (Cell Signaling Technology #7076), donkey anti-mouse Alexa Fluor 488, anti-rabbit Alexa Fluor 594 and anti-sheep Alexa 488 Fluor (ThermoFisher, A-21202, A-21207, and A-11015).…”

Section: Methodsmentioning

confidence: 99%

Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites

et al. 2020

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There are thousands of known cellular phosphorylation sites, but the paucity of ways to identify kinases for particular phosphorylation events remains a major roadblock for understanding kinase signaling. To address this, we here develop a generally applicable method that exploits the large number of kinase inhibitors that have been profiled on near-kinomewide panels of protein kinases. The inhibition profile for each kinase provides a fingerprint that allows identification of unknown kinases acting on target phosphosites in cell extracts. We validate the method on diverse known kinase-phosphosite pairs, including histone kinases, EGFR autophosphorylation, and Integrin β1 phosphorylation by Src-family kinases. We also use our approach to identify the previously unknown kinases responsible for phosphorylation of INCENP at a site within a commonly phosphorylated motif in mitosis (a non-canonical target of Cyclin B-Cdk1), and of BCL9L at S915 (PKA). We show that the method has clear advantages over in silico and genetic screening.

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“…We also identified upregulated (1.684-fold) phosphorylation at the Ser 135 site of histone H 2 B in the yellow sectors (Figure 9E). Previous research indicated that the phosphorylation of H 2 B at Ser6 contributed to chromosomal stability [60]. These phosphoproteins involved in transcriptional regulation might play an important role in leaf color.…”

Section: Discussionmentioning

confidence: 99%

Quantitative Phosphoproteomic and Physiological Analyses Provide Insights into the Formation of the Variegated Leaf in Catalpa fargesii

Wang

Zhu

et al. 2019

IJMS

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Variegated plants are valuable materials for investigating leaf color regulated mechanisms. To unveil the role of posttranslational modification in the variegated phenotype, we conducted global quantitative phosphoproteomic analysis on different leaf color sectors of Maiyuanjinqiu and the corresponding of Catalpa fargesii using Ti4+-IMAC phosphopeptide enrichment. A total of 3778 phosphorylated sites assigned to 1646 phosphoproteins were identified, and 3221 in 1434 proteins were quantified. Differential phosphoproteins (above 1.5 or below 1/1.5) in various leaf color sectors were selected for functional enrichment analyses. Gene ontology (GO) enrichment revealed that processes of photosynthesis, regulation of the generation of precursor metabolites, response to stress, homeostasis, amino acid metabolism, transport–related processes, and most of the energy metabolisms might contribute to leaf color. KEGG pathway enrichment analysis was performed based on differential phosphoproteins (DPs) in different organelles. The result showed that most enriched pathways were located in the chloroplasts and cytosol. The phosphorylation levels of glycometabolism enzymes might greatly affect leaf variegation. Measurements of fluorescence parameters and enzyme activities confirmed that protein phosphorylation could affect plant physiology by regulating enzyme activity. These results provide new clues for further study the formation mechanisms of naturally variegated phenotype.

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CDK1-mediated phosphorylation at H2B serine 6 is required for mitotic chromosome segregation

Cited by 24 publications

References 72 publications

SET binding to Sgo1 inhibits Sgo1–cohesin interactions and promotes chromosome segregation

SET binding to Sgo1 inhibits Sgo1–cohesin interactions and promotes chromosome segregation

Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites

Quantitative Phosphoproteomic and Physiological Analyses Provide Insights into the Formation of the Variegated Leaf in Catalpa fargesii

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