Browse
Submit Data
Databases
API
Help

Dataset Information

21 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Phosphorylation of MCPH1 isoforms during mitosis followed by isoform-specific degradation by APC/C-CDH1.

ABSTRACT: Microcephalin-1 (MCPH1) exists as 2 isoforms that regulate cyclin-dependent kinase-1 activation and chromosome condensation during mitosis, with MCPH1 mutations causing primary microcephaly. MCPH1 is also a tumor suppressor protein, with roles in DNA damage repair/checkpoints. Despite these important roles, there is little information on the cellular regulation of MCPH1. We show that both MCPH1 isoforms are phosphorylated in a cyclin-dependent kinase-1-dependent manner in mitosis and identify several novel phosphorylation sites. Upon mitotic exit, MCPH1 isoforms were degraded by the anaphase-promoting complex/cyclosome-CDH1 E3 ligase complex. Anaphase-promoting complex/cyclosome-CDH1 target proteins generally have D-Box or KEN-Box degron sequences. We found that MCPH1 isoforms are degraded independently, with the long isoform degradation being D-Box dependent, whereas the short isoform was KEN-Box dependent. Our research identifies several novel mechanisms regulating MCPH1 and also highlights important issues with several commercial MCPH1 antibodies, with potential relevance to previously published data.-Meyer, S. K., Dunn, M., Vidler, D. S., Porter, A., Blain, P. G., Jowsey, P. A. Phosphorylation of MCPH1 isoforms during mitosis followed by isoform-specific degradation by APC/C-CDH1.

SUBMITTER: Meyer SK

PROVIDER: S-EPMC6338662 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Phosphorylation and dephosphorylation regulate APC/C(Cdh1) substrate degradation.

Project description:The Anaphase Promoting Complex/Cyclosome (APC/C) ubiquitin ligase activated by its G1 specific adaptor protein Cdh1 is a major regulator of the cell cycle. The APC/C(Cdh1) mediates degradation of dozens of proteins, however, the kinetics and requirements for their degradation are largely unknown. We demonstrate that overexpression of the constitutive active CDH1(m11) mutant that is not inhibited by phosphorylation results in mitotic exit in the absence of the FEAR and MEN pathways, and DNA re-replication in the absence of Cdc7 activity. This mode of mitotic exit also reveals additional requirements for APC/C(Cdh1) substrate degradation, which for some substrates such as Pds1 or Clb5 is dephosphorylation, but for others such as Cdc5 is phosphorylation.

| S-EPMC4825543 | biostudies-literature

APC(cdh1) mediates degradation of the oncogenic Rho-GEF Ect2 after mitosis.

Project description:BACKGROUND: Besides regulation of actin cytoskeleton-dependent functions, Rho GTPase pathways are essential to cell cycle progression and cell division. Rho, Rac and Cdc42 regulate G1 to S phase progression and are involved in cytokinesis. RhoA GDP/GTP cycling is required for normal cytokinesis and recent reports have shown that the exchange factor Ect2 and the GTPase activating protein MgcRacGAP regulate RhoA activity during mitosis. We previously showed that the transcription factors E2F1 and CUX1 regulate expression of MgcRacGAP and Ect2 as cells enter S-phase. METHODOLOGY/PRINCIPAL FINDINGS: We now report that Ect2 is subject to proteasomal degradation after mitosis, following ubiquitination by the APC/C complex and its co-activator Cdh1. A proper nuclear localization of Ect2 is necessary for its degradation. APC-Cdh1 assembles K11-linked poly-ubiquitin chains on Ect2, depending upon a stretch of ?25 amino acid residues that contain a bi-partite NLS, a conventional D-box and two TEK-like boxes. Site-directed mutagenesis of target sequences generated stabilized Ect2 proteins. Furthermore, such degradation-resistant mutants of Ect2 were found to activate RhoA and subsequent signalling pathways and are able to transform NIH3T3 cells. CONCLUSIONS/SIGNIFICANCE: Our results identify Ect2 as a bona fide cell cycle-regulated protein and suggest that its ubiquitination-dependent degradation may play an important role in RhoA regulation at the time of mitosis. Our findings raise the possibility that the overexpression of Ect2 that has been reported in some human tumors might result not only from deregulated transcription, but also from impaired degradation.

| S-EPMC3158779 | biostudies-literature

Phosphorylation-triggered CUEDC2 degradation promotes UV-induced G1 arrest through APC/C(Cdh1) regulation.

Project description:DNA damage triggers cell cycle arrest to provide a time window for DNA repair. Failure of arrest could lead to genomic instability and tumorigenesis. DNA damage-induced G1 arrest is generally achieved by the accumulation of Cyclin-dependent kinase inhibitor 1 (p21). However, p21 is degraded and does not play a role in UV-induced G1 arrest. The mechanism of UV-induced G1 arrest thus remains elusive. Here, we have identified a critical role for CUE domain-containing protein 2 (CUEDC2) in this process. CUEDC2 binds to and inhibits anaphase-promoting complex/cyclosome-Cdh1 (APC/C(Cdh1)), a critical ubiquitin ligase in G1 phase, thereby stabilizing Cyclin A and promoting G1-S transition. In response to UV irradiation, CUEDC2 undergoes ERK1/2-dependent phosphorylation and ubiquitin-dependent degradation, leading to APC/C(Cdh1)-mediated Cyclin A destruction, Cyclin-dependent kinase 2 inactivation, and G1 arrest. A nonphosphorylatable CUEDC2 mutant is resistant to UV-induced degradation. Expression of this stable mutant effectively overrides UV-induced G1-S block. These results establish CUEDC2 as an APC/C(Cdh1) inhibitor and indicate that regulated CUEDC2 degradation is critical for UV-induced G1 arrest.

| S-EPMC3703968 | biostudies-literature

Cyclin E overexpression impairs progression through mitosis by inhibiting APC(Cdh1).

Project description:Overexpression of cyclin E, an activator of cyclin-dependent kinase 2, has been linked to human cancer. In cell culture models, the forced expression of cyclin E leads to aneuploidy and polyploidy, which is consistent with a direct role of cyclin E overexpression in tumorigenesis. In this study, we show that the overexpression of cyclin E has a direct effect on progression through the latter stages of mitotic prometaphase before the complete alignment of chromosomes at the metaphase plate. In some cases, such cells fail to divide chromosomes, resulting in polyploidy. In others, cells proceed to anaphase without the complete alignment of chromosomes. These phenotypes can be explained by an ability of overexpressed cyclin E to inhibit residual anaphase-promoting complex (APC(Cdh1)) activity that persists as cells progress up to and through the early stages of mitosis, resulting in the abnormal accumulation of APC(Cdh1) substrates as cells enter mitosis. We further show that the accumulation of securin and cyclin B1 can account for the cyclin E-mediated mitotic phenotype.

| S-EPMC2064850 | biostudies-literature

Phosphorylation of Skp2 regulated by CDK2 and Cdc14B protects it from degradation by APC(Cdh1) in G1 phase.

Project description:The p27(Kip1) ubiquitin ligase receptor Skp2 is often overexpressed in human tumours and displays oncogenic properties. The activity of SCF(Skp2) is regulated by the APC(Cdh1), which targets Skp2 for degradation. Here we show that Skp2 phosphorylation on Ser64/Ser72 positively regulates its function in vivo. Phosphorylation of Ser64, and to a lesser extent Ser72, stabilizes Skp2 by interfering with its association with Cdh1, without affecting intrinsic ligase activity. Cyclin-dependent kinase (CDK)2-mediated phosphorylation of Skp2 on Ser64 allows its expression in mid-G1 phase, even in the presence of active APC(Cdh1). Reciprocally, dephosphorylation of Skp2 by the mitotic phosphatase Cdc14B at the M --> G1 transition promotes its degradation by APC(Cdh1). Importantly, lowering the levels of Cdc14B accelerates cell cycle progression from mitosis to S phase in an Skp2-dependent manner, demonstrating epistatic relationship of Cdc14B and Skp2 in the regulation of G1 length. Thus, our results reveal that reversible phosphorylation plays a key role in the timing of Skp2 expression in the cell cycle.

| S-EPMC2262036 | biostudies-literature

Isoform-specific monoubiquitination, endocytosis, and degradation of alternatively spliced ErbB4 isoforms.

Project description:Endocytosis and subsequent lysosomal degradation serve as a well characterized mechanism to fine-tune and down-regulate EGFR signaling. However, other members of the EGFR/ErbB receptor family have been reported to be endocytosis-impaired. Here we demonstrate that endocytosis of ErbB4 is regulated in an isoform-specific manner: CYT-1 isoforms were efficiently endocytosed whereas CYT-2 isoforms were endocytosis-impaired. CYT-1 isoforms in endocytic vesicles colocalized with Rab5 and Rab7 indicating trafficking via early endosomes to late endosomal/lysosomal structures. A PPXY motif within the CYT-1-specific sequence that lacks from CYT-2 was necessary both for ubiquitination and endocytosis of CYT-1 isoforms and provided a binding site for a WW domain-containing ubiquitin ligase Itch. Itch catalyzed ubiquitination of ErbB4 CYT-1, promoted its localization into intracellular vesicles, and stimulated degradation of ErbB4 CYT-1. Dominant negative Itch suppressed ErbB4 CYT-1 endocytosis and degradation. These data indicate that ErbB4 isoforms differ in endocytosis and degradation by a mechanism mediated by CYT-1-specific PPXY motif interacting with a WW domain-containing E3 ubiquitin ligase.

| S-EPMC2393780 | biostudies-literature

The spindle checkpoint, APC/C(Cdc20), and APC/C(Cdh1) play distinct roles in connecting mitosis to S phase.

Project description:DNA replication depends on a preceding licensing event by Cdt1 and Cdc6. In animal cells, relicensing after S phase but before mitosis is prevented by the Cdt1 inhibitor geminin and mitotic cyclin activity. Here, we show that geminin, like cyclin B1 and securin, is a bona fide target of the spindle checkpoint and APC/C(Cdc20). Cyclin B1 and geminin are degraded simultaneously during metaphase, which directs Cdt1 accumulation on segregating sister chromatids. Subsequent activation of APC/C(Cdh1) leads to degradation of Cdc6 well before Cdt1 becomes unstable in a replication-coupled manner. In mitosis, the spindle checkpoint supports Cdt1 accumulation, which promotes S phase onset. We conclude that the spindle checkpoint, APC/C(Cdc20), and APC/C(Cdh1) act successively to ensure that the disappearance of licensing inhibitors coincides exactly with a peak of Cdt1 and Cdc6. Whereas cell cycle entry from quiescence requires Cdc6 resynthesis, our results indicate that proliferating cells use a window of time in mitosis, before Cdc6 is degraded, as an earlier opportunity to direct S phase.

| S-EPMC3691463 | biostudies-literature

Differential expression, localization and activity of two alternatively spliced isoforms of human APC regulator CDH1.

Project description:The timely destruction of key regulators through ubiquitin-mediated proteolysis ensures the orderly progression of the cell cycle. The APC (anaphase-promoting complex) is a major component of this degradation machinery and its activation is required for the execution of critical events. Recent studies have just begun to reveal the complex control of the APC through a regulatory network involving WD40 repeat proteins CDC20 and CDH1. In the present paper, we report on the identification and characterization of human CDH1beta, a novel alternatively spliced isoform of CDH1. Both CDH1alpha and CDH1beta can bind to the APC and stimulate the degradation of cyclin B1, but they are differentially expressed in human tissues and cells. CDH1alpha contains a nuclear localization signal which is absent in CDH1beta. Intracellularly, CDH1alpha appears in the nucleus whereas CDH1beta is a predominantly cytoplasmic protein. The forced overexpression of CDH1alpha in cultured cells correlates with the reduction of nuclear cyclin A, but the steady-state amount of cyclin A does not change noticeably in CDH1beta-overexpressed cells. In Xenopus embryos, ectopic overexpression of human CDH1alpha, but not of CDH1beta, induces cell-cycle arrest during the first G(1) phase at the mid-blastula transition. Taken together, our findings document the differential expression, subcellular localization and cell-cycle-regulatory activity of human CDH1 isoforms.

| S-EPMC1223613 | biostudies-other

The E3 ligase APC/C-Cdh1 regulates MEF2A-dependent transcription by targeting SUMO-specific protease 2 for ubiquitination and degradation.

Project description:Activity-dependent stimuli induced a calcineurin-mediated dephosphorylation of the transcriptional factor MEF2A at serine408 and promoted a switch from SUMOylation to acetylation at lysine403 which led to MEF2A transcriptional activation. We previously identified SENP2 is the de-SUMOylation enzyme for MEF2A and promotes MEF2A-dependent transcription. We report here a requirement for APC(Cdh1)-SENP2-MEF2A axis in the regulation of MEF2A transcriptional activation. APC(Cdh1) interacts with and targets SENP2 for ubiquitination and destruction in the cytoplasm by recognizing a conserved canonical D-box motif in SENP2. Moreover, Cdh1 regulates the transcriptional activity of MEF2A in a SENP2 dependent manner. Activity-dependent stimuli prevented APC(Cdh1)-induced SENP2 ubiquitination, promoted SENP2 nuclear accumulations, and caused MEF2A de-SUMOylation and MEF2A acetylation, leading to MEF2A transcriptional activation. Thus, our findings defined a post-transcriptional mechanism underlying activity-dependent stimuli-induced MEF2A transcriptional activation.

| S-EPMC4612642 | biostudies-literature

Multiple mechanisms determine the order of APC/C substrate degradation in mitosis.

Project description:The ubiquitin protein ligase anaphase-promoting complex or cyclosome (APC/C) controls mitosis by promoting ordered degradation of securin, cyclins, and other proteins. The mechanisms underlying the timing of APC/C substrate degradation are poorly understood. We explored these mechanisms using quantitative fluorescence microscopy of GFP-tagged APC/C(Cdc20) substrates in living budding yeast cells. Degradation of the S cyclin, Clb5, begins early in mitosis, followed 6 min later by the degradation of securin and Dbf4. Anaphase begins when less than half of securin is degraded. The spindle assembly checkpoint delays the onset of Clb5 degradation but does not influence securin degradation. Early Clb5 degradation depends on its interaction with the Cdk1-Cks1 complex and the presence of a Cdc20-binding "ABBA motif" in its N-terminal region. The degradation of securin and Dbf4 is delayed by Cdk1-dependent phosphorylation near their Cdc20-binding sites. Thus, a remarkably diverse array of mechanisms generates robust ordering of APC/C(Cdc20) substrate destruction.

| S-EPMC4195823 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data