Phosphate-binding pocket on cyclin B governs CDK substrate phosphorylation and mitotic timing - NCOMMS-24-17810-T - 2
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Orbitrap Fusion Lumos
ORGANISM(S): Saccharomyces Cerevisiae (ncbitaxon:4932)
SUBMITTER: 
Raymond Suhandynata
PROVIDER: MSV000094550 | MassIVE | Tue Apr 16 00:55:00 BST 2024
SECONDARY ACCESSION(S): PXD051477
REPOSITORIES: MassIVE
Similar Datasets
Project description:Cell cycle progression is governed by complexes of the cyclin-dependent kinases (CDKs) and their regulatory subunits cyclin and Cks1. CDKs phosphorylate hundreds of substrates, often at multiple sites. Multisite phosphorylation depends on Cks1, which binds initial priming phosphorylation sites to promote secondary phosphorylation at other sites. Here, we describe a similar role for a recently discovered phosphate-binding pocket (PP) on B-type cyclins. Mutation of the PP in Clb2, the major mitotic cyclin of budding yeast, alters bud morphology and delays the onset of anaphase. Using phosphoproteomics in vivo and kinase reactions in vitro, we find that mutation of the PP reduces phosphorylation of several CDK substrates, including the Bud6 subunit of the polarisome and the Cdc16 and Cdc27 subunits of the anaphase-promoting complex/cyclosome. We conclude that the cyclin PP, like Cks1, controls the timing of multisite phosphorylation on CDK substrates, thereby helping to establish the robust timing of cell-cycle events.
| S-EPMC10925351 | biostudies-literature
Project description:We have repurposed standard phosphoproteomics workflow based on Fe3+IMAC for enrichment ofmannose-6-phosphate modified glycopeptides. This worklfow was used to profile lysosomal acid hydrolases in HeLa and CHO cell lines. We have combined this approach with CRISPR/Cas9 KO of acid phosphatases 2 and 5 responsible for dephosphorylation of mannose-6-phosphate glycopeptides in the lysosome. This combined approach enabled significantly deeper coverage of CHO mannose-6-phosphate glycoproteome.
2018-12-18 | PXD010333 | Pride
Project description:Background The establishment of tissue architecture requires coordination between distinct processes including basement membrane assembly, cell adhesion, and polarity; however, the underlying mechanisms remain poorly understood. The actin cytoskeleton is ideally situated to orchestrate tissue morphogenesis due to its roles in mechanical, structural, and regulatory processes. However, the function of many pivotal actin-binding proteins in mammalian development is poorly understood. Results Here, we identify a crucial role for anillin (ANLN), an actin-binding protein, in orchestrating epidermal morphogenesis. In utero RNAi-mediated silencing of Anln in mouse embryos disrupted epidermal architecture marked by adhesion, polarity, and basement membrane defects. Unexpectedly, these defects cannot explain the profoundly perturbed epidermis of Anln-depleted embryos. Indeed, even before these defects emerge, Anln-depleted epidermis exhibits abnormalities in mitotic rounding and its associated processes: chromosome segregation, spindle orientation, and mitotic progression, though not in cytokinesis that was disrupted only in Anln-depleted cultured keratinocytes. We further show that ANLN localizes to the cell cortex during mitotic rounding, where it regulates the distribution of active RhoA and the levels, activity, and structural organization of the cortical actomyosin proteins. Conclusions Our results demonstrate that ANLN is a major regulator of epidermal morphogenesis and identify a novel role for ANLN in mitotic rounding, a near-universal process that governs cell shape, fate, and tissue morphogenesis. Supplementary Information The online version contains supplementary material available at 10.1186/s12915-022-01345-9.
| S-EPMC9205045 | biostudies-literature
Project description:Post-mitotic cell separation is one of the most prominent events in the life cycle of eukaryotic cells, but the molecular underpinning of this fundamental biological process is far from being concluded and fully characterized. We use budding yeast Saccharomyces cerevisiae as a model and demonstrate AMN1 as a major gene underlying post-mitotic cell separation in a natural yeast strain, YL1C. Specifically, we define a novel 11-residue domain by which Amn1 binds to Ace2. Moreover, we demonstrate that Amn1 induces proteolysis of Ace2 through the ubiquitin proteasome system and in turn, down-regulates Ace2's downstream target genes involved in hydrolysis of the primary septum, thus leading to inhibition of cell separation and clumping of haploid yeast cells. Using ChIP assays and site-specific mutation experiments, we show that Ste12 and the a1-α12 heterodimer are two direct regulators of AMN1. Specifically, a1-α2, a diploid-specific heterodimer, prevents Ste12 from inactivating AMN1 through binding to its promoter. This demonstrates how the Amn1-governed cell separation is highly cell type dependent. Finally, we show that AMN1368D from YL1C is a dominant allele in most strains of S. cerevisiae and evolutionarily conserved in both genic structure and phenotypic effect in two closely related yeast species, K. lactis and C. glabrata.
| S-EPMC6181423 | biostudies-literature
Project description:The faithful segregation of intact genetic material and the perpetuation of chromatin states through mitotic cell divisions are pivotal for maintaining cell function and identity across cell generations. However, most exogenous mutagens generate long-lasting DNA lesions that are segregated during mitosis. How this segregation is controlled is unknown. Here, we uncover a mitotic chromatin-marking pathway that governs the segregation of UV-induced damage in human cells. Our mechanistic analyses reveal two layers of control: histone ADP-ribosylation, and the incorporation of newly synthesized histones at UV damage sites, that both prevent local mitotic phosphorylations on histone H3 serines. Functionally, this chromatin-marking pathway drives the asymmetric segregation of UV damage in the cell progeny with potential consequences on daughter cell fate. We propose that this mechanism may help preserve the integrity of stem cell compartments during asymmetric cell divisions.
| S-EPMC10508772 | biostudies-literature
Project description:Accurate control of the Ras-related nuclear protein (Ran) GTPase cycle depends on the regulated activity of regulator of chromosome condensation 1 (RCC1), Ran's nucleotide exchange factor. RanBP1 has been characterized as a coactivator of the Ran GTPase-activating protein RanGAP1. RanBP1 can also form a stable complex with Ran and RCC1, although the dynamics and function of this complex remain poorly understood. Here, we show that formation of the heterotrimeric RCC1/Ran/RanBP1 complex in M phase Xenopus egg extracts controls both RCC1's enzymatic activity and partitioning between the chromatin-bound and soluble pools of RCC1. This mechanism is critical for spatial control of Ran-guanosine triphosphate (GTP) gradients that guide mitotic spindle assembly. Moreover, phosphorylation of RanBP1 drives changes in the dynamics of chromatin-bound RCC1 pools at the metaphase-anaphase transition. Our findings reveal an important mitotic role for RanBP1, controlling the spatial distribution and magnitude of mitotic Ran-GTP production and thereby ensuring accurate execution of Ran-dependent mitotic events.
| S-EPMC4254504 | biostudies-literature
Project description:Nano-flow LC-MS/MS analysis of 293T digest in two gradients.
2023-08-01 | PXD044221 |
Project description:Polyporus umbellatus sclerotia are medicinal tissue, but little is known about acetylome of sclerotia generated from hyphae. Based on acetylome analysis results and expression levels of targeted protein that associated with sclerotia generation were verified by PRM using Orbitrap Fusion (Thermo-Fisher Scientific, San Jose, CA) mass spectrometer equipped with a nanospray Flex Ion Source and the UltiMate3000 RSLC nano (Dionex, Sunnyvale, CA).
2022-05-20 | PXD031198 | Pride
Project description:Phosphoproteome of Fe3+ IMAC flowthrough from enriched human mitotic spindles
2006-02-24 | PRD000007 | Pride
Project description:Fe-IMAC columns for robust and reproducible phosphopeptide ernichment, comparison to TiO2 batch and Ti-IMAC tip enrichment, large scale phosphoproteomics coupling Fe-IMAC column pre-enrichment to subsequent hSAX separation
2015-08-19 | PXD001060 | Pride