Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Rho family of GTPases plays key roles in the regulation of cell motility and morphogenesis. They also regulate protein kinase cascades, gene expression, and cell cycle progression. This multiplicity of roles requires that the Rho GTPases interact with a wide variety of downstream effector proteins. An understanding of their functions at a molecular level therefore requires the identification of the entire set of such effectors. Towards this end, we performed a two-hybrid screen using the TC10 GTPase as bait and identified a family of putative effector proteins related to MSE55, a murine stromal and epithelial cell protein of 55 kDa. We have named this family the Borg (binder of Rho GTPases) proteins. Complete open reading frames have been obtained for Borg1 through Borg3. We renamed MSE55 as Borg5. Borg1, Borg2, Borg4, and Borg5 bind both TC10 and Cdc42 in a GTP-dependent manner. Surprisingly, Borg3 bound only to Cdc42. An intact CRIB (Cdc42, Rac interactive binding) domain was required for binding. No interaction of the Borgs with Rac1 or RhoA was detectable. Three-hemagglutinin epitope (HA(3))-tagged Borg3 protein was mostly cytosolic when expressed ectopically in NIH 3T3 cells, with some accumulation in membrane ruffles. The phenotype induced by Borg3 was reminiscent of that caused by an inhibition of Rho function and was reversed by overexpression of Rho. Surprisingly, it was independent of the ability to bind Cdc42. Borg3 also inhibited Jun kinase activity by a mechanism that was independent of Cdc42 binding. HA(3)-Borg3 expression caused substantial delays in the spreading of cells on fibronectin surfaces after replating, and the spread cells lacked stress fibers. We propose that the Borg proteins function as negative regulators of Rho GTPase signaling.
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PMID:The Borgs, a new family of Cdc42 and TC10 GTPase-interacting proteins. 1049 May 98

Rho-associated kinase (Rho-kinase) is a serine/threonine protein kinase downstream of the small GTPase Rho, which participates in signaling pathways of many cellular functions. Although Rho-kinase is implicated in the regulation of the morphology of neuronal cells, the distribution of Rho-kinase in the brain has not been elucidated yet. In this study, we investigated the distribution of Rho-kinase using three antibodies recognizing the different epitopes of Rho-kinase. Rho-kinase was abundantly expressed in the gray matter in comparison with the white matter. Strong immunoreactivity was observed in the pyramidal neurons of the cerebral cortex and hippocampus and in the Purkinje cells of the cerebellum. These results indicate that Rho-kinase is abundantly distributed in neurons and might play an important role in remodeling of neurites.
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PMID:Distribution of Rho-kinase in the bovine brain. 1049 34

Dendritogenesis, axonogenesis, pathfinding, and target recognition are all affected in distinct ways when Xenopus retinal ganglion cells (RGCs) are transfected with constitutively active (ca), wild-type (wt), and dominant negative (dn) Rho-family GTPases in vivo. Dendritogenesis required Rac1 and Cdc42 activity. Moreover, ca-Rac1 caused dendrite hyperproliferation. Axonogenesis, in contrast, was inhibited by ca-Rac1. This phenotype was partially rescued by the coexpression of dn cyclin-dependent kinase (Cdk5), a proposed effector of Rac1, suggesting that Rac1 activity must be regulated tightly for normal axonogenesis. Growth cone morphology was particularly sensitive to dn-RhoA and wt-Cdc42 constructs. These also caused targeting errors, such as tectal bypass, suggesting that cytoskeletal rearrangements are involved in target recognition and are transduced by these pathways.
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PMID:The neuronal architecture of Xenopus retinal ganglion cells is sculpted by rho-family GTPases in vivo. 1049 46

Platelets undergo shape change upon activation with agonists. During shape change, disc-shaped platelets turn into spiculated spheres with protruding filopodia. When agonist-induced cytosolic Ca(2+) increases were prevented using the cytosolic Ca(2+) chelator, 5, 5'-dimethyl-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (5, 5'-dimethyl-BAPTA), platelets still underwent shape change, although the onset was delayed and the initial rate was dramatically decreased. In the absence of cytosolic Ca(2+), agonist-stimulated myosin light chain phosphorylation was significantly inhibited. The myosin light chain was maximally phosphorylated at 2 s in control platelets compared with 30 s in 5,5'-dimethyl-BAPTA-treated platelets. ADP, thrombin, or U46619-induced Ca(2+)-independent platelet shape change was significantly reduced by staurosporine, a nonselective kinase inhibitor, by the selective p160 Rho-associated coiled-coil-containing protein kinase inhibitor Y-27632, or by HA 1077. Both Y-27632 and HA 1077 reduced peak levels of ADP-induced platelet shape change and myosin light chain phosphorylation in control platelets. In 5,5'-dimethyl-BAPTA-treated platelets, Y-27632 and HA 1077 completely abolished both ADP-induced platelet shape change and myosin light chain phosphorylation. Our results indicate that Ca(2+)/calmodulin-stimulated myosin light chain kinase and p160 Rho-associated coiled-coil-containing protein kinase independently contribute to myosin light chain phosphorylation and platelet shape change, through Ca(2+)-sensitive and Ca(2+)-insensitive pathways, respectively.
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PMID:Platelet shape change is mediated by both calcium-dependent and -independent signaling pathways. Role of p160 Rho-associated coiled-coil-containing protein kinase in platelet shape change. 1049 86

Human hepatocellular carcinoma (HCC) can invade the portal vein and metastasizes to other parts of the liver even at a relatively early stage of the disease, with less tumor spread occurring outside the liver. This intrahepatic metastasis is the main cause of liver failure and death in HCC patients. To analyze the mechanisms of intrahepatic metastasis we have constructed metastatic models using orthotopic implantation of human HCC cell lines. Five HCC cell lines formed liver tumors after injection into the livers of SCID mice, and of those 5 cell lines, Li7 and KYN-2 cells also resulted in vascular tumor thrombi and intrahepatic metastasis. These 2 cell lines had markedly higher cell motilities than the other 3 cell lines in vitro. Their motilities appeared to be Rho-mediated; serum and lysophosphatidic acid (LPA) evoked actin reorganization and motility of Li7 cells, and C3 exoenzyme exposure reduced the motility of both serum-stimulated Li7 cells and KYN-2 cells. Dominant negative and active forms of p160 Rho-associated coiled-coil forming protein kinase (p160ROCK), one of the downstream effectors of Rho, were separately and stably introduced into Li7 cells. Dominant active p160ROCK transfectants showed increased motility that was independent of serum and LPA, and dominant negative p160ROCK transfectants showed reduced motility under stimulation. Furthermore, implantation of dominant negative p160ROCK transfectants resulted in a reduced metastatic rate in vivo compared with the parent cells or a control transfectant. These findings indicate that cell motility mediated by the Rho/p160ROCK signaling pathway plays a critical role in intrahepatic metastasis of human HCC.
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PMID:Cell motility mediated by rho and Rho-associated protein kinase plays a critical role in intrahepatic metastasis of human hepatocellular carcinoma. 1049 56

Rho-kinase is implicated in the phosphorylation of myosin light chain downstream of Rho, which is thought to induce smooth muscle contraction and stress fiber formation in non-muscle cells. Here, we examined the mode of action of inhibitors of Rho-kinase. The chemical compounds such as HA1077 and Y-32885 inhibited not only the Rho-kinase activity but also the activity of protein kinase N, one of the targets of Rho, but had less of an effect on the activity of myotonic dystrophy kinase-related Cdc42-binding kinase beta (MRCKbeta). The COOH-terminal portion of Rho-kinase containing Rho-binding (RB) and pleckstrin homology (PH) domains (RB/PH (TT)), in which point mutations were introduced to abolish the Rho binding activity, interacted with Rho-kinase and thereby inhibited the Rho-kinase activity, whereas RB/PH (TT) had no effect on the activity of protein kinase N or MRCKbeta, suggesting that the COOH-terminal region of Rho-kinase is a possible negative regulatory region of Rho-kinase. The expression of RB/PH (TT) specifically blocked the stress fiber and focal adhesion formation induced by the active form of Rho or Rho-kinase in NIH 3T3 cells, but not that induced by the active form of MRCKbeta or myosin light chain. Thus, RB/PH (TT) appears to specifically inhibit Rho-kinase in vivo.
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PMID:The COOH terminus of Rho-kinase negatively regulates rho-kinase activity. 1054 85

Ras proteins are guanine nucleotide-binding proteins that play pivotal roles in the control of normal and transformed cell growth and are among the most intensively studied proteins of the past decade. After stimulation by various growth factors and cytokines, Ras activates several downstream effectors, including the Raf-1/mitogen-activated protein kinase pathway and the Rac/Rho pathway. In approximately 30% of human cancers, including a substantial proportion of pancreatic and colon adenocarcinomas, mutated ras genes produce mutated proteins that remain locked in an active state, thereby relaying uncontrolled proliferative signals. Ras undergoes several posttranslational modifications that facilitate its attachment to the inner surface of the plasma membrane. The first-and most critical-modification is the addition of a farnesyl isoprenoid moiety in a reaction catalyzed by the enzyme protein farnesyltransferase (FTase). It follows that inhibiting FTase would prevent Ras from maturing into its biologically active form, and FTase is of considerable interest as a potential therapeutic target. Different classes of FTase inhibitors have been identified that block farnesylation of Ras, reverse Ras-mediated cell transformation in human cell lines, and inhibit the growth of human tumor cells in nude mice. In transgenic mice with established tumors, FTase inhibitors cause regression in some tumors, which appears to be mediated through both apoptosis and cell cycle regulation. FTase inhibitors have been well tolerated in animal studies and do not produce the generalized cytotoxic effects in normal tissues that are a major limitation of most conventional anticancer agents. There are ongoing clinical evaluations of FTase inhibitors to determine the feasibility of administering them on dose schedules like those that portend optimal therapeutic indices in preclinical studies. Because of the unique biologic aspects of FTase, designing disease-directed phase II and III evaluations of their effectiveness presents formidable challenges.
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PMID:Ras protein farnesyltransferase: A strategic target for anticancer therapeutic development. 1055 Jan 63

1 G protein-mediated Ca2+ sensitization of airway smooth muscle contraction was investigated with respect to the relative importance of Rho-associated coiled coil forming protein kinase (ROCK) and protein kinase C (PKC). We examined the effects of Y-27632, a ROCK inhibitor, and GF 109203X, a PKC inhibitor, on guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS)-induced contraction in alpha-toxin- or beta-escin-permeabilized rabbit trachea. 2 Although pre-treatment with Y-27632 dose-dependently inhibited GTPgammaS (10 microM)-induced Ca2+ sensitization of alpha-toxin-permeabilized trachea, a Y-27632-insensitive component (approximately 16% of the maximum contraction) was retained during the early phase of the GTPgammaS response in the presence of Y-27632 (100 microM). 3 GF 109203X (5 microM) abolished 1 microM 4beta-phorbol 12, 13-dibutyrate (PDBu)-induced, but only partially inhibited the GTPgammaS-induced Ca2+ sensitization. A combination of Y-27632 (100 microM) and GF 109203X (5 microM) totally abolished the GTPgammaS response. 4 GTPgammaS caused only a small contraction in the absence of Ca2+. Wortmannin (30 microM), a myosin light chain kinase (MLCK) inhibitor, completely inhibited Ca2+-induced contraction. ATP-triggered contraction of the strip which had been treated with calyculin A (1 microM), a phosphatase inhibitor, in rigor solutions was markedly slowed by worthmannin (30 microM), but not by Y-27632 (100 microM), in the presence of GTPgammaS and Ca2+. 5 GTPgammaS, but not PDBu, contracted the beta-escin-permeabilized trachea in the absence of Ca2+, but the presence of Ca2+-independent MLCK. 6 We conclude that ROCK plays a primary role in G-protein-mediated Ca2+ sensitization, which requires MLCK activity, with minor contribution of PKC to the early phase of contraction, and PDBu utilizes conventional PKC(s) in airway smooth muscle.
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PMID:A major role for the rho-associated coiled coil forming protein kinase in G-protein-mediated Ca2+ sensitization through inhibition of myosin phosphatase in rabbit trachea. 1055 27

The small GTPase Rho; functions as a molecular switch that regulates various cellular processes such as cell adhesion, motility, gene expression and cytokinesis. We previously isolated several putative Rho; targets including rhophilin which bound selectively to the GTP-bound form of Rho;. Rhophilin is expressed highly in testis and is localized specifically in sperm flagella. The presence of a PDZ domain at the carboxy terminus of rhophilin suggested that rhophilin works as an adaptor molecule. To test this hypothesis, we employed a yeast two hybrid system using the rhophilin PDZ domain as a bait, and screened a mouse testis cDNA library. We isolated several positive clones containing the same insert. The open reading frame of the cDNA encoded a novel protein of 212 amino acids designated as ropporin from a Japanese word 'oppo' (the tail). The amino-terminal 40 amino acid sequence of ropporin showed high homology to that of the regulatory subunit of type II cAMP-dependent protein kinase, which is involved in dimerization and binding to A-kinase anchoring proteins. Consistently, a yeast two hybrid assay and gel filtration of recombinant ropporin indicated that ropporin dimerizes through this domain. Deletion analysis indicated that the carboxy-terminal four amino acids are essential for binding of ropporin to rhophilin, and ropporin and RhoV14 coprecipitated in the presence of rhophilin in vitro. Northern blot analysis showed that ropporin is exclusively expressed in testis, and induced at the late stage of spermatogenesis. This induction paralleled that of rhophilin. Immunocytochemistry using anti-ropporin antibody showed that ropporin is localized in the principal piece and the end piece of sperm flagella. Electronmicroscopy revealed that ropporin is mostly localized in the inner surface of the fibrous sheath while rhophilin is present in the outer surface of the outer dense fiber. These results suggest that rhophilin and ropporin may form a complex in sperm flagella.
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PMID:Ropporin, a sperm-specific binding protein of rhophilin, that is localized in the fibrous sheath of sperm flagella. 1059 29

The bioactive phospholipid lysophosphatidic acid (LPA) causes growth cone collapse and neurite retraction in neuronal cells. These changes are brought about by the action of a cell surface receptor coupled to specific G proteins that control morphology and motility through the action of a group of small GTPases, the Rho family of proteins. Many studies have focused on actin reorganization modulated by Rho-GTPases, but almost no information has been obtained concerning microtubular network reorganization after LPA-induced neurite retraction. In the present study, we demonstrate an increase in site-specific Alzheimer's disease-like Tau phosphorylation during LPA-induced neurite retraction in differentiated SY-SH5Y human neuroblastoma cells. The phosphorylation state of Tau was inferred from its immunoreactivity with antibodies that recognize phosphorylation-sensitive epitopes. The effects of specific kinase inhibitors indicate that this phosphorylation is mediated by glycogen synthase kinase-3 (GSK-3). In support of this idea, we observed an increase of GSK-3 activity upon growth cone collapse. Our results are consistent with the hypothesis that activation of GSK-3 occurs in the Rho pathway and may represent an important link between microtubules and microfilaments dynamics during neuritogenesis and in pathological situations such as Alzheimer's disease.
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PMID:The neurite retraction induced by lysophosphatidic acid increases Alzheimer's disease-like Tau phosphorylation. 1060 Dec 62


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