Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cyclin-dependent kinase Cdc28p associates with the cyclin Clb2p to induce mitosis in the yeast Saccharomyces cerevisiae. Several cell cycle regulatory proteins have been shown to require specific nuclear transport events to exert their regulatory functions. Therefore, we investigated the subcellular localization of wild-type Clb2p and several mutant versions of the protein using green fluorescent protein (GFP) fusion constructs. Wild-type Clb2p is primarily nuclear at all points of the cell. A point mutation in a potential leucine-rich nuclear export signal (NES) enhances the nuclear localization of the protein, and delta-yrb2 cells exhibit an apparent Clb2p nuclear export defect. Clb2p contains a bipartite nuclear localization signal (NLS), and its nuclear localization requires the alpha and beta importins (Srp1p and Kap95p), as well as the yeast Ran GTPase and its regulators. Deletion of the Clb2p NLS causes increased cytoplasmic localization of the protein, as well as accumulation at the bud neck. These data indicate that Clb2p exists in multiple places in the yeast cell, possibly allowing Cdc28p to locally phosphorylate substrates at distinct subcellular sites.
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PMID:The Saccharomyces cerevisiae cyclin Clb2p is targeted to multiple subcellular locations by cis- and trans-acting determinants. 1117 27

Different G(1) cyclins confer functional specificity to the cyclin-dependent kinase (Cdk) Cdc28p in budding yeast. The Cln3p G(1) cyclin is localized primarily to the nucleus, while Cln2p is localized primarily to the cytoplasm. Both binding to Cdc28p and Cdc28p-dependent phosphorylation in the C-terminal region of Cln2p are independently required for efficient nuclear depletion of Cln2p, suggesting that this process may be physiologically regulated. The accumulation of hypophosphorylated Cln2 in the nucleus is an energy-dependent process, but may not involve the RAN GTPase. Phosphorylation of Cln2p is inefficient in small newborn cells obtained by elutriation, and this lowered phosphorylation correlates with reduced Cln2p nuclear depletion in newborn cells. Thus, Cln2p may have a brief period of nuclear residence early in the cell cycle. In contrast, the nuclear localization pattern of Cln3p is not influenced by Cdk activity. Cln3p localization requires a bipartite nuclear localization signal (NLS) located at the C terminus of the protein. This sequence is required for nuclear localization of Cln3p and is sufficient to confer nuclear localization to green fluorescent protein in a RAN-dependent manner. Mislocalized Cln3p, lacking the NLS, is much less active in genetic assays specific for Cln3p, but more active in assays normally specific for Cln2p, consistent with the idea that Cln3p localization explains a significant part of Clnp functional specificity.
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PMID:Mechanisms controlling subcellular localization of the G(1) cyclins Cln2p and Cln3p in budding yeast. 1150 71

The Saccharomyces cerevisiae morphogenesis checkpoint delays mitosis in response to insults that impair actin organization and/or bud formation. The delay is due to accumulation of the inhibitory kinase Swe1p, which phosphorylates the cyclin-dependent kinase Cdc28p. Having screened through a panel of yeast mutants with defects in cell morphogenesis, we report here that the polarity establishment protein Bem2p is required for the checkpoint response. Bem2p is a Rho-GTPase activating protein (GAP) previously shown to act on Rho1p, and we now show that it also acts on Cdc42p, the GTPase primarily responsible for establishment of cell polarity in yeast. Whereas the morphogenesis role of Bem2p required GAP activity, the checkpoint role of Bem2p did not. Instead, this function required an N-terminal Bem2p domain. Thus, this single protein has a GAP-dependent role in promoting cell polarity and a GAP-independent role in responding to defects in cell polarity by enacting the checkpoint. Surprisingly, Swe1p accumulation occurred normally in bem2 cells, but they were nevertheless unable to promote Cdc28p phosphorylation. Therefore, Bem2p defines a novel pathway in the morphogenesis checkpoint.
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PMID:The Rho-GAP Bem2p plays a GAP-independent role in the morphogenesis checkpoint. 1214 2

The Rho family GTPase Cdc42 is recognized for its role in cellular proliferation and transformation. However, the mechanism by which it promotes cell cycle progression has remained undefined. Using an inducible expression system, we show that constitutively active Cdc42 (Cdc42V12) is sufficient by itself to induce anchorage-independent but not mitogen-independent growth in NIH3T3 cells. However, Cdc42V12 markedly accelerates activation of cyclin E-Cdk2 in response to mitogen. These effects were highly specific, as the kinetics of cyclin D-Cdk4 activation was unaltered. Cdc42V12 promotes Cdk2 activation by selectively inducing cyclin E expression without affecting other regulatory proteins such as the p27 Cdk inhibitor or Cdc25A. Furthermore, Cdc42V12 was able to activate a reporter gene driven by the cyclin E promoter in the absence of exogenous mitogen or adhesion. Cyclin E induction was sensitive to rapamycin but not inhibitors of mitogen-activated protein kinases, implicating p70 S6 kinase (p70S6k) as the relevant mediator. Consistent with this notion, wild type and constitutively active alleles of p70S6k were sufficient to activate the cyclin E promoter. In sum, these studies provide novel insights into the mechanism by which Cdc42 promotes G1 progression.
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PMID:Cdc42 promotes G1 progression through p70 S6 kinase-mediated induction of cyclin E expression. 1284 76

In this study we have used the transgenic mouse model Tg2576 to analyze the involvement of anomalous loss of regulation of cdk5 and the stress kinases JNK and p38 in brain neuronal death as related to neurodegenerative disorders such as Alzheimer's disease. Previous studies on hippocampal cells led us to the discovery that the cdk5/p35 complex is activated in neurodegeneration, a finding that was confirmed later in the transgenic mouse model. Here we show a link between the cdk5 system and JNK and p38 phosphoproteins, as an alternative pathway to neuronal death. Brains of the Tg2576 transgenic mice overexpressing amyloid precursor protein exhibited immunoreactivity with the phosphoproteins p-JNK, p-p38 and the GTPase protein Rac1 surrounding neuritic plaques. A significant increase in the immunodetection of JNK and p38 phosphoproteins in the Tg2576 mouse compared with wild type controls confirmed these findings. The significant increase in co-immunoprecipitation of p-JNK, p-p38 and Rac1 proteins with cdk5 in the transgenic mouse provided evidence for these interactions. At the cellular level, p-JNK and cdk5 colocalized in the cytoplasm of the cell bodies and neurites of brain cortical areas of the transgenic mouse. The present evidence suggests a cellular link between the cdk5 system and the stress kinase JNK and p38 pathways in an in vivo model. This study sheds new light on the pathogenesis of neuronal degeneration processes such as those occurring in Alzheimer's disease.
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PMID:Modulation of the JNK and p38 pathways by cdk5 protein kinase in a transgenic mouse model of Alzheimer's disease. 1466

How proliferating cells maintain the copy number and overall size of their organelles is not clear. We had previously reported that in the budding yeast Saccharomyces cerevisiae the G1 cyclin Cln3p is required for vacuolar (lysosomal) homotypic fusion and loss of Cln3p leads to vacuolar fragmentation. The Cdc42p GTPase is also required for vacuole fusion. Here we show that the scaffold protein Bem1p, a critical regulator of Cdc42p activity, is a downstream effector of Cln3p and the cyclin-dependent kinase (Cdk) Cdc28p. Our results suggest that Bem1p is phosphorylated in a Cdk-dependent manner to promote vacuole fusion. Replacing Ser72 with Asp, to mimic phosphorylation at an optimal Cdk-consensus site located in the first SH3 domain of Bem1p, suppressed vacuolar fragmentation in cells lacking Cln3p. Using in vivo and in vitro assays, we found that Cln3p was unable to promote vacuole fusion in the absence of Bem1p or in the presence of a nonphosphorylatable Bem1p-Ser72Ala mutant. Furthermore, activation of Cdc42p also suppressed vacuolar fragmentation in the absence of Cln3p. Our results provide a mechanism that links cyclin-dependent kinase activity with vacuole fusion through Bem1p and the Cdc42p GTPase cycle.
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PMID:Bem1p, a scaffold signaling protein, mediates cyclin-dependent control of vacuolar homeostasis in Saccharomyces cerevisiae. 1623 May 27

We have identified two redundant GTPase activating proteins (GAPs) - RGA-3 and RGA-4 - that regulate Rho GTPase function at the plasma membrane in early Caenorhabditis elegans embryos. Knockdown of both RhoGAPs resulted in extensive membrane ruffling, furrowing and pronounced pseudo-cleavages. In addition, the non-muscle myosin NMY-2 and RHO-1 accumulated on the cortex at sites of ruffling. RGA-3 and RGA-4 are GAPs for RHO-1, but most probably not for CDC-42, because only RHO-1 was epistatic to the two GAPs, and the GAPs had no obvious influence on CDC-42 function. Furthermore, knockdown of either the RHO-1 effector, LET-502, or the exchange factor for RHO-1, ECT-2, alleviated the membrane-ruffling phenotype caused by simultaneous knockdown of both RGA-3 and RGA-4 [rga-3/4 (RNAi)]. GFP::PAR-6 and GFP::PAR-2 were localized at the anterior and posterior part of the early C. elegans embryo, respectively showing that rga-3/4 (RNAi) did not interfere with polarity establishment. Most importantly, upon simultaneous knockdown of RGA-3, RGA-4 and the third RhoGAP present in the early embryo, CYK-4, NMY-2 spread over the entire cortex and GFP::PAR-2 localization at the posterior cortex was greatly diminished. These results indicate that the functions of CYK-4 are temporally and spatially distinct from RGA-3 and RGA-4 (RGA-3/4). RGA-3/4 and CYK-4 also play different roles in controlling LET-502 activation in the germ line, because rga-3/4 (RNAi), but not cyk-4 (RNAi), aggravated the let-502(sb106) phenotype. We propose that RGA-3/4 and CYK-4 control with which effector molecules RHO-1 interacts at particular sites at the cortex in the zygote and in the germ line.
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PMID:Functions of the novel RhoGAP proteins RGA-3 and RGA-4 in the germ line and in the early embryo of C. elegans. 1772 51

Cyclin-dependent kinases (CDKs) trigger essential cell cycle processes including critical events in G1 phase that culminate in bud emergence, spindle pole body duplication, and DNA replication. Localized activation of the Rho-type GTPase Cdc42p is crucial for establishment of cell polarity during G1, but CDK targets that link the Cdc42p module with cell growth and cell cycle commitment have remained largely elusive. Here, we identify the GTPase-activating protein (GAP) Rga2p as an important substrate related to the cell polarity function of G1 CDKs. Overexpression of RGA2 in the absence of functional Pho85p or Cdc28p CDK complexes is toxic, due to an inability to polarize growth. Mutation of CDK consensus sites in Rga2p that are phosphorylated both in vivo and in vitro by Pho85p and Cdc28p CDKs results in a loss of G1 phase-specific phosphorylation. A failure to phosphorylate Rga2p leads to defects in localization and impaired polarized growth, in a manner dependent on Rga2p GAP function. Taken together, our data suggest that CDK-dependent phosphorylation restrains Rga2p activity to ensure appropriate activation of Cdc42p during cell polarity establishment. Inhibition of GAPs by CDK phosphorylation may be a general mechanism to promote proper G1-phase progression.
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PMID:Activation of the Cdc42p GTPase by cyclin-dependent protein kinases in budding yeast. 1785 95

Site-specific activation of the Rho-type GTPase Cdc42p is critical for the establishment of cell polarity. Here we investigated the role and regulation of the GTPase-activating enzymes (GAPs) Bem2p and Bem3p for Cdc42p activation and actin polarization at bud emergence in Saccharomyces cerevisiae. Bem2p and Bem3p are localized throughout the cytoplasm and the cell cortex in unbudded G1 cells, but accumulate at sites of polarization after bud emergence. Inactivation of Bem2p results in hyperactivation of Cdc42p and polarization toward multiple sites. Bem2p and Bem3p are hyperphosphorylated at bud emergence most likely by the Cdc28p-Cln2p kinase. This phosphorylation appears to inhibit their GAP activity in vivo, as non-phosphorylatable Bem3p mutants are hyperactive and interfere with Cdc42p activation. Taken together, our results indicate that Bem2p and Bem3p may function as global inhibitors of Cdc42p activation during G1, and their inactivation by the Cdc28p/Cln kinase contributes to site-specific activation of Cdc42p at bud emergence.
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PMID:Phosphorylation of Bem2p and Bem3p may contribute to local activation of Cdc42p at bud emergence. 1791 57

RCC1, a guanine nucleotide exchange factor for Ran GTPase, plays essential roles in the growth and viability of mammalian cells. Here, we examined the phosphorylation of specific serine and threonine residues of RCC1 in vivo and showed that RCC1 is indeed phosphorylated. Analysis by two-dimensional (2D) gel electrophoresis suggested that serine 11 (S11) of hamster RCC1 is phosphorylated in vivo. A point mutation of S11 of hamster RCC1 resulted in a decrease in the number of 2D gel spots, indicating a lack of phosphorylation at the mutant residue. S11 phosphorylation in vitro depended on cyclin B-cdc2 kinase. An RCC1 mutant in which all N-terminal serine and threonine residues were substituted with glutamate residues to mimic phosphorylation at these residues showed decreased binding to the karyopherin, KPNA4, compared with wild type RCC1. We conclude that RCC1 undergoes post-translational phosphorylation.
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PMID:Ran GTPase guanine nucleotide exchange factor RCC1 is phosphorylated on serine 11 by cdc2 kinase in vitro. 1856 22


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