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

Mitosis-specific phosphorylation by cdc2 kinase causes nonmuscle caldesmon to dissociate from microfilaments (Yamashiro, S., Yamakita, Y., Ishikawa, R., and Matsumura, F. (1990) Nature 344, 675-678; Yamashiro, S., Yamakita, Y., Hosoya, H., and Matsumura, F. (1991) Nature 349, 169-172). To explore the function of mitosis-specific phosphorylation of caldesmon, in vivo- and in vitro-phosphorylated caldesmons have been characterized. We have found that both in vivo and in vitro phosphorylation of caldesmon causes similar changes in the properties, including reduction in actin, calmodulin, and myosin binding of caldesmon, and a decrease in the inhibition of actomyosin ATPase by caldesmon. Rat non-muscle caldesmon is phosphorylated in vitro up to a ratio of 7 mol/mol of protein. Actin-binding constants of both a high affinity (K a = 1.2 x 10(7) M-1) and a low affinity (K a = 1 x 10(6) M-1) site of unphosphorylated caldesmon are reduced to less than 10(5) M-1 with 5 mol of phosphate incorporation per mol of protein. Actin-bound caldesmon can be phosphorylated by cdc2 kinase, which results in the dissociation of caldesmon from F-actin. Caldesmon has a second myosin-binding site in the C terminus, in addition to the N terminus myosin-binding domain previously reported, because the bacterially expressed C terminus of caldesmon shows binding to myosin. Phosphorylation of the C-terminal fragments decreases their myosin-binding affinity as observed with intact caldesmon. These results suggest that caldesmon loses most of its in vitro functions during mitosis as a result of phosphorylation, which may be required for the reorganization of microfilaments during mitosis.
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PMID:Characterization of mitotically phosphorylated caldesmon. 153 4

One of the profound changes in cellular morphology which occurs during mitosis is a massive alteration in the organization of the microfilament cytoskeleton. This change, together with other mitotic events including nuclear membrane breakdown, chromosome condensation and formation of mitotic spindles, is induced by a molecular complex called maturation promoting factor. This consists of at least two subunits, a polypeptide of relative molecular mass 45,000-62,000 (Mr 45-62K) known as cyclin, and a 34K catalytic subunit which has serine/threonine kinase activity and is known as cdc2 kinase. Non-muscle caldesmon, an 83K actin- and calmodulin-binding protein, is dissociated from microfilaments during mitosis, apparently as a consequence of mitosis-specific phosphorylation. We now report that cdc2 kinase phosphorylates caldesmon in vitro principally at the same sites as those phosphorylated in vivo during mitosis, and that phosphorylation reduces the binding affinity of caldesmon for both actin and calmodulin. Because caldesmon inhibits actomyosin ATPase, our results suggest that cdc2 kinase directly causes microfilament reorganization during mitosis.
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PMID:Phosphorylation of non-muscle caldesmon by p34cdc2 kinase during mitosis. 198 9

Phosphorylation of rat non-muscle caldesmon by cdc2 kinase causes reduction in most of caldesmon's properties, including caldesmon's binding to actin, myosin, and calmodulin, as well as its inhibition of actomyosin ATPase. We have generated and characterized the COOH terminus of caldesmon mutants lacking mitosis-specific phosphorylation sites, because the COOH-terminal half of caldesmon contains all 7 putative Ser or Thr sites for cdc2 kinase. Codons for the 7 putative Ser or Thr residues have been mutated to Ala, and resultant mutants were bacterially expressed. Analyses of the phosphopeptide maps of these mutants have identified 6 sites, including Ser-249, Ser-462, Thr-468, Ser-491, Ser-497, and Ser-527 as the mitosis-specific phosphorylation sites, whereas the phosphorylation of the remaining site, Thr-377, is not detected by this assay method. Actin binding experiments have suggested that 5 sites including Ser-249, Ser-462, Thr-468, Ser-491, and Ser-497 are important for the phosphorylation-dependent reduction in actin binding. Characterization of a mutant lacking all 7 Ser or Thr sites (7-fold mutant) has revealed that 7-fold mutation eliminates all phosphorylation sites by cdc2 kinase. While the in vitro properties of the 7-fold mutant, including actin, myosin, and calmodulin binding and inhibition of actomyosin ATPase, are very similar to those of nonmutated protein, such properties are not affected by the treatment with cdc2 kinase in contrast to nonmutated protein. This mutant should thus be useful to explore the functions of the mitosis-specific phosphorylation of caldesmon.
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PMID:Characterization of the COOH terminus of non-muscle caldesmon mutants lacking mitosis-specific phosphorylation sites. 787 50

Mitosis-specific phosphorylation by cdc2 kinase causes nonmuscle caldesmon to dissociate from microfilaments during prometaphase. (Yamashiro, S., Y. Yamakita, R. Ishikawa, and F. Matsumura. 1990. Nature (Lond.). 344:675-678; Yamashiro, S., Y. Yamakita, H. Hosoya, and F. Matsumura. 1991. Nature (Lond.) 349:169-172). To explore the functions of caldesmon phosphorylation during cytokinesis, we have examined the relationship between the phosphorylation level, actin-binding, and in vivo localization of caldesmon in cultured cells after their release of metaphase arrest. Immunofluorescence studies have revealed that caldesmon is localized diffusely throughout cytoplasm in metaphase. During early stages of cytokinesis, caldesmon is still diffusely present and not concentrated in contractile rings, in contrast to the accumulation of actin in cleavage furrows during cytokinesis. In later stages of cytokinesis, most caldesmon is observed to be yet diffusely localized although some concentration of caldesmon is observed in cortexes as well as in cleavage furrows. When daughter cells begin to spread, caldesmon shows complete colocalization with F-actin-containing structures. These observations are consistent with changes in the levels of microfilament-associated caldesmon during synchronized cell division. Caldesmon is missing from microfilaments in prometaphase cells arrested by nocodazole treatment, as shown previously (Yamashiro, S., Y. Yamakita, R. Iskikawa, and F. Matsumura. 1990. Nature (Lond.). 344:675-678). The level of microfilament-associated caldesmon stays low (12% of that of interphase cells) when some cells start cytokinesis at 40 min after the release of metaphase arrest. When 60% of cells finish cytokinesis at 60 min, the level of microfilament-associated caldesmon is recovered to 50% of that of interphase cells. The level of microfilament-associated caldesmon is then gradually increased to 80% when cells show spreading at 120 min. Dephosphorylation appears to occur during cytokinesis. It starts when cells begin to show cytokinesis at 40 min and completes when most cells finish cytokinesis at 60 min. These results suggest that caldesmon is not associated with microfilaments of cleavage furrows at least in initial stages of cytokinesis and that dephosphorylation of caldesmon appears to couple with its reassociation with microfilaments. Because caldesmon is known to inhibit actomyosin ATPase and/or regulate actin assembly, its continued dissociation from microfilaments may be required for the assembly and/or activation of contractile rings.
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PMID:Localization of caldesmon and its dephosphorylation during cell division. 838 77

The mitotic disassembly and reassembly of the mammalian Golgi apparatus is an ideal system to study the molecular mechanisms involved in biogenesis and maintenance of membranous organelles. As cells enter M-phase, Golgi stacks are converted into Golgi clusters of small membrane fragments, which are dispersed throughout the cytoplasmic space during metaphase. Disassembly is dependent on the action of cdc2-kinase and at least two distinct pathways contribute to the fragmentation: one involves the budding of COP I-coated vesicles from Golgi cisternae, the other is a less well characterised COP I-independent pathway. During telophase, the Golgi fragments reassemble and fuse into a fully functional Golgi stack, using at least two distinct ATPase-mediated fusion pathways.
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PMID:Molecular mechanisms in the disassembly and reassembly of the mammalian Golgi apparatus during M-phase. 868 8

Transcription factor IIH (TFIIH) is a multisubunit complex required for transcription and for DNA nucleotide excision repair. TFIIH possesses three enzymatic activities: (i) an ATP-dependent DNA helicase, (ii) a DNA-dependent ATPase, and (iii) a kinase with specificity for the carboxyl-terminal domain of RNA polymerase II. The kinase activity was recently identified as the cdk (cyclin-dependent kinase) activating kinase, CAK, composed of cdk7, cyclin H, and MAT-1. Here we report the isolation and characterization of three distinct CAK-containing complexes from HeLa nuclear extracts: CAK, a novel CAK-ERCC2 complex, and TFIIH. CAK-ERCC2 can efficiently associate with core-TFIIH to reconstitute holo-TFIIH transcription activity. We present evidence proposing a critical role for ERCC2 in mediating the association of CAK with core TFIIH subunits.
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PMID:Human cyclin-dependent kinase-activating kinase exists in three distinct complexes. 869 42

The transcription/DNA repair factor TFIIH consists of nine subunits, several exhibiting known functions: helicase/ATPase, kinase activity and DNA binding. Three subunits of TFIIH, cdk7, cyclin H and MAT1, form a ternary complex, cdk-activating kinase (CAK), found either on its own or as part of TFIIH. In the present work, we demonstrate that purified human CAK complex (free CAK) and recombinant CAK (rCAK) produced in insect cells exhibit a strong preference for the cyclin-dependent kinase 2 (cdk2) over a ctd oligopeptide substrate (which mimics the carboxy-terminal domain of the RNA polymerase II). In contrast, TFIIH preferentially phosphorylates the ctd as well as TFIIE alpha, but not cdk2. TFIIH was resolved into four subcomplexes: the kinase complex composed of cdk7, cyclin H and MAT1; the core TFIIH which contains XPB, p62, p52, p44 and p34; and two other subcomplexes in which XPD is found associated with either the kinase complex or with the core TFIIH. Using these fractions, we demonstrate that TFIIH lacking the CAK subcomplex completely recovers its transcriptional activity in the presence of free CAK. Furthermore, studies examining the interactions between TFIIH subunits provide evidence that CAK is integrated within TFIIH via XPB and XPD.
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PMID:Substrate specificity of the cdk-activating kinase (CAK) is altered upon association with TFIIH. 913 Jul 8

Nucleolin is a major protein of exponentially growing eukaryotic cells where it is present in abundance at the heart of the nucleolus. It is highly conserved during evolution. Nucleolin contains a specific bipartite nuclear localization signal sequence and possesses a number of unusual structural features. It has unique tripartite structure and each domain performs a specific function by interacting with DNA or RNA or proteins. Nucleolin exhibits intrinsic self-cleaving, DNA helicase, RNA helicase and DNA-dependent ATPase activities. Nucleolin also acts as a sequence-specific RNA binding protein, an autoantigen, and as the component of a B cell specific transcription factor. Its phosphorylation by cdc2, CK2, and PKC-zeta modulate some of its activities. This multifunctional protein has been implicated to be involved directly or indirectly in many metabolic processes such as ribosome biogenesis (which includes rDNA transcription, pre-rRNA synthesis, rRNA processing, ribosomal assembly and maturation), cytokinesis, nucleogenesis, cell proliferation and growth, cytoplasmic-nucleolar transport of ribosomal components, transcriptional repression, replication, signal transduction, inducing chromatin decondensation and many more (see text). In plants it is developmentally, cell-cycle, and light regulated. The regulation of all these functions of a single protein seems to be a challenging puzzle.
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PMID:Nucleolin: a multifunctional major nucleolar phosphoprotein. 991 13

The pathogenesis of diabetic neuropathy remains unclear, although several factors have been implicated in its pathogenesis. We have examined possible roles of decreased production of nitric oxide, ion channel dysfunction and decreased capacity of nerve regeneration. STZ-induced diabetic rats showed decreases in nociceptive threshold and NADPH-diaphorase positive neurons, nNOS level and cGMP content of DRG at 12 weeks after induction of diabetes. The rats injected by L-NAME, potent nNOS inhibitor, showed decreased nociceptive threshold, although D-NAME, inactive in nNOS inhibition, did not. These results suggest that decreased NO production might be involved in hyperalgesia in diabetic rats. Both hyperglycemia and decreased Na/K-ATPase activity are thought to be characteristic features of diabetic neuropathy. To investigate the presence of ion channel abnormality in diabetic nerves, a Vaseline-gap voltage clamp technique was applied for a single myelinated fibers under 30 mM high glucose plus 0.1 mM ouabain. Since K current was increased, a Ca activated K channel blocker was applied and this increase was shown to be suppressed. Furthermore, Ca channel blockers all suppressed increased K currents, suggesting that the condition induced an increase of Ca influx, thereby increasing Ca activated K currents through K channels. The data are important in that diabetic condition may induce both Ca influx, leading to nerve degeneration, and increased K current, resulting in decreased nerve conduction. Nerve regeneration has been known to be disturbed in diabetic condition. We have shown a decrease in nerve elongation rate in diabetic rats after crush of sciatic nerve, although this decrease was not ameliorated by ARI. Furthermore, Wallerian degeneration was shown to be delayed in diabetic nerves, leading to delayed nerve regeneration. Hyperphosphorylation of both medium and high molecular weight neurofilaments that might be induced by protein kinases including CDK 5 may be involved in the mechanism.
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PMID:[New trend in pathogenesis of diabetic neuropathy]. 1037 17

Cells require optimum protein synthetic activity in order to support cell proliferation, maintain homeostatic and metabolic integrity, and repair damage. Since growth depends on protein synthesis through ribosome biogenesis, the control of biosynthesis of ribosomes is necessarily a key element for control of growth. Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation. The highly conserved nucleolin contains three major domains through which it controls the organization of nucleolar chromatin, packaging of pre-RNA, rDNA transcription, and ribosome assembly. Numerous reports have implicated the involvement of nucleolin either directly or indirectly in the regulation of cell proliferation and growth, cytokinesis, replication, embryogenesis, and nucleogenesis. Nucleolin, an RNA binding protein, is also an autoantigen, a transcriptional repressor, and a switch region targeting factor. In addition, nucleolin exhibits autodegradation, DNA and RNA helicase activities, and DNA-dependent ATPase activity. An interesting aspect of nucleolin action is that it is a target for regulation by proteolysis, methylation, ADP-ribosylation, and phosphorylation by CKII, cdc2, PKC-xi, cyclic AMP-dependent protein kinase, and ecto-protein kinase. For these and other reasons, nucleolin is fundamental to the survival and proliferation of cells. Considerable progress has been made in recent years with the identification of new nucleolin binding proteins that may mediate these many nucleolin-dependent functions. Nucleolin also functions as a cell surface receptor, where it acts as a shuttling protein between cytoplasm and nucleus, and thus can even provide a mechanism for extracellular regulation of nuclear events. Exploration of the regulation of this multifaceted protein in a remarkable number of diverse functions is challenging.
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PMID:Molecular dissection of nucleolin's role in growth and cell proliferation: new insights. 1054 74


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