The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Yoshimi Takai - One of the best experts on this subject based on the ideXlab platform.
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rac P21 is involved in insulin induced membrane ruffling and rho P21 is involved in hepatocyte growth factor and 12 o tetradecanoylphorbol 13 acetate tpa induced membrane ruffling in kb cells
Molecular and Cellular Biology, 1994Co-Authors: Takayuki Nishiyama, Kenji Takaishi, Takuya Sasaki, Masaki Kato, H Yaku, K Araki, Y Matsuura, Yoshimi TakaiAbstract:Insulin and hepatocyte growth factor (HGF) induced morphologically different membrane rufflings in KB cells. Insulin-induced membrane ruffling was inhibited by microinjection of rho GDI, an inhibitory GDP/GTP exchange regulator for both rho P21 and rac P21 small GTP-binding proteins, but not inhibited by microinjection of botulinum exoenzyme C3, known to selectively ADP-ribosylate rho P21 and to impair its function. This rho GDI action was prevented by comicroinjection with guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-bound rac1 P21. In contrast, HGF-induced membrane ruffling was inhibited by microinjection of rho GDI or C3. This rho GDI action was prevented by comicroinjection with GTP gamma S-bound rhoA P21, and this C3 action was prevented by comicroinjection with GTP gamma S-bound rhoAIle-41 P21, which is resistant to C3. Microinjection of either GTP gamma S-bound rac1 P21 or rhoA P21 alone induced membrane ruffling in the absence of the growth factors. The rac1 P21-induced membrane ruffling was morphologically similar to the insulin-induced kind, whereas rhoA P21-induced ruffling was apparently different from both the insulin- and HGF-induced kinds. Membrane ruffling was also induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), a protein kinase C-activating phorbol ester, but not by Ca2+ ionophore or microinjection of a dominant active Ki-ras P21 mutant (Ki-rasVal-12 P21). The phorbol ester-induced membrane ruffling was morphologically similar to the rhoA P21-induced kind and inhibited by microinjection of rho GDI or C3. These results indicate that rac P21 and rho GDI are involved in insulin-induced membrane ruffling and that rho P21 and rho GDI are involved in HGF- and phorbol ester-induced membrane rufflings.
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involvement of rho P21 small gtp binding protein and its regulator in the hgf induced cell motility
Oncogene, 1994Co-Authors: Kenji Takaishi, Takuya Sasaki, Masaki Kato, Wataru Yamochi, Shinya Kuroda, T Nakamura, Masatoshi Takeichi, Yoshimi TakaiAbstract:Hepatocyte growth factor (HGF) induced motility of cultured mouse keratinocytes (308R cells). This HGF-induced cell motility was inhibited by microinjection of either rho GDI, an inhibitory GDP/GTP exchange protein for rho P21 small GTP-binding protein, or a botulinum exoenzyme C3 which is known to selectively impair the function of rho P21 by ADP-ribosylating its effector domain. The rho GDI action was prevented by comicroinjection with the guanosine 5'-(3-0-thio)triphosphate (GTP gamma S)-bound active form of rhoA P21, and the C3 action was prevented by comicroinjection with a rhoA P21 mutant (rhoAIle41 P21) which is resistant to the C3 action. The HGF-induced cell motility was not inhibited by microinjection of a dominant negative rac1 P21 mutant (rac1Asn17 P21) or a dominant negative Ki-ras P21 mutant (Ki-rasAsn17 P21). Microinjection of the GTP gamma S-bound form of rac1 P21 or a dominant active Ki-ras P21 mutant (Ki-rasVal12 P21) did not induce cell motility. These results indicate that both rho P21 and rho GDI, but neither rac P21 nor ras P21, are involved in the HGF-induced cell motility. However, microinjection of the GTP gamma S-bound form of rhoA P21 alone did not induce cell motility in the absence of HGF, suggesting that activation of rho P21 is necessary but not sufficient for the HGF-induced cell motility. The HGF-induced cell motility was mimicked by 12-0-tetradecanoyl-phorbol-13-acetate, a protein kinase C-activating phorbol ester, but not by Ca2+ ionophore. The phorbol ester-induced cell motility was also inhibited by microinjection of rho GDI or C3. These results indicate that both rho P21 and rho GDI are also involved in the phorbol ester-induced cell motility.
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regulation of morphology by rho P21 and its inhibitory gdp gtp exchange protein rho gdi in swiss 3t3 cells
Journal of Biological Chemistry, 1993Co-Authors: Yasushi Miura, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, H Yaku, T Musha, Yoshimi TakaiAbstract:rho GDP dissociation inhibitor (GDI) is an inhibitory GDP/GTP exchange protein for a group of small GTP-binding proteins including at least rhoA P21, rhoB P21, rac1 P21, rac2 P21, and G25K. Microinjection of rho GDI into Swiss 3T3 cells made the cells round and refractile. This morphological change was accompanied by the disappearance of stress fibers. The rho GDI action was prevented by comicroinjection of rho GDI with the guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-bound form of rhoA P21, but not with the GTP gamma S-bound form of rhoA P21 lacking the C-terminal three amino acids, which was not post-translationally modified with lipids. The GTP gamma S-bound form of rac1 P21, the same form of G25K, the same form of smg P21B, or Ki-rasval12 P21 was ineffective. Microinjection of the bacterial ADP-ribosyltransferase C3 specific for rho P21 into Swiss 3T3 cells induced the similar changes of morphology and stress fibers. This C3 action was not prevented by comicroinjection of C3 with the GTP gamma S-bound form of rhoA P21, but was prevented by comicroinjection with the same form of a rhoA P21 mutant which was not ADP-ribosylated by C3. These results indicate that the rho GDI-rho P21 system regulates cell morphology presumably through the actomyosin system in Swiss 3T3 cells.
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involvement of rho P21 and its inhibitory gdp gtp exchange protein rho gdi in cell motility
Molecular and Cellular Biology, 1993Co-Authors: Kenji Takaishi, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, Ko Kotani, Yoshimi TakaiAbstract:Evidence is accumulating that rho P21, a ras P21-related small GTP-binding protein (G protein), regulates the actomyosin system. The actomyosin system is known to be essential for cell motility. In the present study, we examined the action of rho P21, its inhibitory GDP/GTP exchange protein (named rho GDI), its stimulatory GDP/GTP exchange protein (named smg GDS), and Clostridium botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho P21 and to impair its function, in cell motility (chemokinesis) of Swiss 3T3 cells. We quantitated the capacity of cell motility by measuring cell tracks by phagokinesis. Microinjection of the GTP gamma S-bound active form of rhoA P21 or smg GDS into Swiss 3T3 cells did not affect cell motility, but microinjection of rho GDI into the cells did inhibit cell motility. This rho GDI action was prevented by comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA P21 but not with the same form of rhoA P21 lacking the C-terminal three amino acids which was not posttranslationally modified with lipids. The rho GDI action was not prevented by Ki-rasVal-12 P21 or any of the GTP gamma S-bound form of other small GTP-binding proteins including rac1 P21, G25K, and smg P21B. Among these small G proteins, rhoA P21, rac1 P21, and G25K are known to be substrates for rho GDI. The rho GDI action was not prevented by comicroinjection of rho GDI with smg GDS. Microinjection of C3 into Swiss 3T3 cells also inhibited cell motility. These results indicate that the rho GDI-rho P21 system regulates cell motility, presumably through the actomyosin system.
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the functional domain of the stimulatory gdp gtp exchange protein smg gds which interacts with the c terminal geranylgeranylated region of rap1 krev 1 smg P21
Oncogene, 1992Co-Authors: K Kotani, Akira Kikuchi, K Doi, Shosei Kishida, Tsuyoshi Sakoda, K Kishi, Yoshimi TakaiAbstract:rap1/Krev-1/smg P21 (smg P21), a member of the small GTP-binding protein (G protein) superfamily, has a geranylgeranylated cysteine residue and clustered basic amino acids in the C-terminal region. The GDP/GTP exchange reaction of smg P21 is regulated by smg GDS, which is also active on Ki-ras P21 and rho P21. The C-terminal region of smg P21 is essential for its interaction with smg GDS. Moreover, smg P21 is phosphorylated by cyclic AMP- and cyclic GMP-dependent protein kinases at the serine residue between the polybasic region and the prenylated cysteine residue, and this phosphorylation initiates the smg GDS-induced smg P21 activation. Thus, the C-terminal cationic and hydrophobic region is important for the regulation of the smg P21 activity. In the present study, we attempted to determine the functional domain of smg GDS which interacts with the C-terminal region of smg P21 by use of a cross-link method and a site-directed mutagenesis method. The region of smg GDS cross-linked with the C-terminal region of smg P21B was residues 444-492, which is located at the C-terminal fifth of smg GDS. On deletion of these residues, smg GDS became inactive on smg P21B, Ki-ras P21 and rhoA P21. These results indicate that residues 444-492 of smg GDS are at least one of the domains which interact with the C-terminal region of its substrate small G proteins.
Akira Kikuchi - One of the best experts on this subject based on the ideXlab platform.
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rap1 P21 regulates the interaction of ras P21 with rgl a new effector protein of ras P21
FEBS Letters, 1995Co-Authors: Masahiro Ikeda, Shinya Koyama, Michiko Okazaki, Kiyohiko Dohi, Akira KikuchiAbstract:We have recently found that ralGDS family members (RGL and ralGDS) are putative effector proteins of ras P21. rap1 P21 is a small GTP-binding protein which has the same amino acid sequence as the effector loop of ras P21. We examined the effect of rap1 P21 on the interaction of ras P21 with RGL. The GTP-bound form of rap1 P21 interacted with RGL as well as did ras P21. rap1 P21 inhibited the interaction of ras P21 with RGL. RGL was phosphorylated by cyclic AMP-dependent protein kinase (protein kinase A). Phosphorylation of RGL did not affect its binding to ras P21 and rap1 P21 under the conditions that phosphorylation of Raf-1 reduced its affinity for ras P21. These results demonstrate that rap1 P21 but not protein kinase A regulates the interaction of ras P21 with RGL and suggest that rap1 P21 and protein kinase A may cooperate to distinguish the signal of ras P21 to RGL from that to Raf-1.
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regulation of morphology by rho P21 and its inhibitory gdp gtp exchange protein rho gdi in swiss 3t3 cells
Journal of Biological Chemistry, 1993Co-Authors: Yasushi Miura, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, H Yaku, T Musha, Yoshimi TakaiAbstract:rho GDP dissociation inhibitor (GDI) is an inhibitory GDP/GTP exchange protein for a group of small GTP-binding proteins including at least rhoA P21, rhoB P21, rac1 P21, rac2 P21, and G25K. Microinjection of rho GDI into Swiss 3T3 cells made the cells round and refractile. This morphological change was accompanied by the disappearance of stress fibers. The rho GDI action was prevented by comicroinjection of rho GDI with the guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-bound form of rhoA P21, but not with the GTP gamma S-bound form of rhoA P21 lacking the C-terminal three amino acids, which was not post-translationally modified with lipids. The GTP gamma S-bound form of rac1 P21, the same form of G25K, the same form of smg P21B, or Ki-rasval12 P21 was ineffective. Microinjection of the bacterial ADP-ribosyltransferase C3 specific for rho P21 into Swiss 3T3 cells induced the similar changes of morphology and stress fibers. This C3 action was not prevented by comicroinjection of C3 with the GTP gamma S-bound form of rhoA P21, but was prevented by comicroinjection with the same form of a rhoA P21 mutant which was not ADP-ribosylated by C3. These results indicate that the rho GDI-rho P21 system regulates cell morphology presumably through the actomyosin system in Swiss 3T3 cells.
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involvement of rho P21 and its inhibitory gdp gtp exchange protein rho gdi in cell motility
Molecular and Cellular Biology, 1993Co-Authors: Kenji Takaishi, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, Ko Kotani, Yoshimi TakaiAbstract:Evidence is accumulating that rho P21, a ras P21-related small GTP-binding protein (G protein), regulates the actomyosin system. The actomyosin system is known to be essential for cell motility. In the present study, we examined the action of rho P21, its inhibitory GDP/GTP exchange protein (named rho GDI), its stimulatory GDP/GTP exchange protein (named smg GDS), and Clostridium botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho P21 and to impair its function, in cell motility (chemokinesis) of Swiss 3T3 cells. We quantitated the capacity of cell motility by measuring cell tracks by phagokinesis. Microinjection of the GTP gamma S-bound active form of rhoA P21 or smg GDS into Swiss 3T3 cells did not affect cell motility, but microinjection of rho GDI into the cells did inhibit cell motility. This rho GDI action was prevented by comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA P21 but not with the same form of rhoA P21 lacking the C-terminal three amino acids which was not posttranslationally modified with lipids. The rho GDI action was not prevented by Ki-rasVal-12 P21 or any of the GTP gamma S-bound form of other small GTP-binding proteins including rac1 P21, G25K, and smg P21B. Among these small G proteins, rhoA P21, rac1 P21, and G25K are known to be substrates for rho GDI. The rho GDI action was not prevented by comicroinjection of rho GDI with smg GDS. Microinjection of C3 into Swiss 3T3 cells also inhibited cell motility. These results indicate that the rho GDI-rho P21 system regulates cell motility, presumably through the actomyosin system.
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the functional domain of the stimulatory gdp gtp exchange protein smg gds which interacts with the c terminal geranylgeranylated region of rap1 krev 1 smg P21
Oncogene, 1992Co-Authors: K Kotani, Akira Kikuchi, K Doi, Shosei Kishida, Tsuyoshi Sakoda, K Kishi, Yoshimi TakaiAbstract:rap1/Krev-1/smg P21 (smg P21), a member of the small GTP-binding protein (G protein) superfamily, has a geranylgeranylated cysteine residue and clustered basic amino acids in the C-terminal region. The GDP/GTP exchange reaction of smg P21 is regulated by smg GDS, which is also active on Ki-ras P21 and rho P21. The C-terminal region of smg P21 is essential for its interaction with smg GDS. Moreover, smg P21 is phosphorylated by cyclic AMP- and cyclic GMP-dependent protein kinases at the serine residue between the polybasic region and the prenylated cysteine residue, and this phosphorylation initiates the smg GDS-induced smg P21 activation. Thus, the C-terminal cationic and hydrophobic region is important for the regulation of the smg P21 activity. In the present study, we attempted to determine the functional domain of smg GDS which interacts with the C-terminal region of smg P21 by use of a cross-link method and a site-directed mutagenesis method. The region of smg GDS cross-linked with the C-terminal region of smg P21B was residues 444-492, which is located at the C-terminal fifth of smg GDS. On deletion of these residues, smg GDS became inactive on smg P21B, Ki-ras P21 and rhoA P21. These results indicate that residues 444-492 of smg GDS are at least one of the domains which interact with the C-terminal region of its substrate small G proteins.
Myriam Gorospe - One of the best experts on this subject based on the ideXlab platform.
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lincrna P21 suppresses target mrna translation
Molecular Cell, 2012Co-Authors: Jehyun Yoon, Xiaoling Yang, Kotb Abdelmohsen, Subramanya Srikantan, Jennifer L Martindale, Maite Huarte, Ming Zhan, Kevin G Becker, Myriam GorospeAbstract:Mammalian long intergenic noncoding RNAs (lincRNAs) are best known for modulating transcription. Here we report a posttranscriptional function for lincRNA-P21 as a modulator of translation. Association of the RNA-binding protein HuR with lincRNA-P21 favored the recruitment of let-7/Ago2 to lincRNA-P21, leading to lower lincRNA-P21 stability. Under reduced HuR levels, lincRNA-P21 accumulated in human cervical carcinoma HeLa cells, increasing its association with JUNB and CTNNB1 mRNAs and selectively lowering their translation. With elevated HuR, lincRNA-P21 levels declined, which in turn derepressed JunB and β-catenin translation and increased the levels of these proteins. We propose that HuR controls translation of a subset of target mRNAs by influencing lincRNA-P21 levels. Our findings uncover a role for lincRNA as a posttranscriptional inhibitor of translation.
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Prostaglandin A2-mediated stabilization of P21 mRNA through an ERK-dependent pathway requiring the RNA-binding protein HuR.
The Journal of biological chemistry, 2004Co-Authors: Xiaoling Yang, Wengong Wang, Jinshui Fan, Ashish Lal, Dongmei Yang, Heping Cheng, Myriam GorospeAbstract:Treatment with the stress agent prostaglandin A2 (PGA2) induces expression of the cyclin-dependent kinase inhibitor P21. Here, we present evidence that P21 expression increases through PGA2-triggered stabilization of the P21 mRNA and further show that these events require the mitogen-activated protein (MAP) kinase ERK. Binding experiments using either endogenous P21 mRNA or in vitro-labeled P21 transcripts revealed a specific PGA2-dependent association of the P21 mRNA with the RNA-binding protein HuR. Interestingly, although inhibition of the ERK pathway did not prevent the PGA2-triggered increase in cytoplasmic HuR, it did impair the formation of endogenous and in vitro [HuR-P21 mRNA] complexes and further prevented the PGA2-mediated stabilization of the P21 mRNA, suggesting that ERK-mediated events were required for binding HuR to the P21 mRNA and preventing its decay. RNA interference-based knockdown of HuR abundance further served to demonstrate the contribution of HuR-mediated P21 mRNA stabilization toward enhancing P21 expression after PGA2 treatment. Collectively, our results indicate that PGA2 stabilizes the P21 mRNA through an ERK-independent increase in cytoplasmic HuR levels and an ERK-dependent association of HuR with the P21 mRNA.
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hur regulates P21 mrna stabilization by uv light
Molecular and Cellular Biology, 2000Co-Authors: Wengong Wang, Henry Furneaux, Huiming Cheng, Craig M Caldwell, Dorothy Hutter, Yusen Liu, Nikki J Holbrook, Myriam GorospeAbstract:Expression of the cyclin-dependent kinase inhibitor P21 is highly induced by many stresses, including exposure to short-wavelength UV light (UVC), which increases P21 mRNA stability. Investigation into the mechanisms underlying this stabilization process revealed that proteins present in cytoplasmic lysates of human RKO colorectal carcinoma cells formed complexes with P21 mRNA that were inducible by treatment with UVC and other stress agents. The ubiquitous Elav-type RNA-binding protein HuR was identified within the P21 mRNA-protein complexes, as antibodies recognizing HuR supershifted these complexes and revealed HuR-immunoreactive proteins complexing with P21 mRNA on Western blots. Lowering of endogenous HuR levels through expression of antisense HuR decreased P21 RNA-protein complexes, greatly reduced the UVC inducibility and half-life of P21 mRNA, and prevented UVC-mediated induction of luciferase activity in P21 3* untranslated region-containing reporter constructs. Our findings indicate that HuR plays a major role in regulating stress-induced P21 expression by enhancing P21 mRNA stability and that these effects are coupled to HuR’s elevated presence in the cytoplasm. Expression of P21, a universal inhibitor of cyclin-dependent kinases, has been found to increase following exposure to a wide variety of stress agents, including genotoxins, oxidants, and metabolic perturbations. Indeed, increased P21 expression is believed to participate in mediating the growth arrest that follows exposure to such insults (15, 24, 62) and affects pro
Takuya Sasaki - One of the best experts on this subject based on the ideXlab platform.
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rac P21 is involved in insulin induced membrane ruffling and rho P21 is involved in hepatocyte growth factor and 12 o tetradecanoylphorbol 13 acetate tpa induced membrane ruffling in kb cells
Molecular and Cellular Biology, 1994Co-Authors: Takayuki Nishiyama, Kenji Takaishi, Takuya Sasaki, Masaki Kato, H Yaku, K Araki, Y Matsuura, Yoshimi TakaiAbstract:Insulin and hepatocyte growth factor (HGF) induced morphologically different membrane rufflings in KB cells. Insulin-induced membrane ruffling was inhibited by microinjection of rho GDI, an inhibitory GDP/GTP exchange regulator for both rho P21 and rac P21 small GTP-binding proteins, but not inhibited by microinjection of botulinum exoenzyme C3, known to selectively ADP-ribosylate rho P21 and to impair its function. This rho GDI action was prevented by comicroinjection with guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-bound rac1 P21. In contrast, HGF-induced membrane ruffling was inhibited by microinjection of rho GDI or C3. This rho GDI action was prevented by comicroinjection with GTP gamma S-bound rhoA P21, and this C3 action was prevented by comicroinjection with GTP gamma S-bound rhoAIle-41 P21, which is resistant to C3. Microinjection of either GTP gamma S-bound rac1 P21 or rhoA P21 alone induced membrane ruffling in the absence of the growth factors. The rac1 P21-induced membrane ruffling was morphologically similar to the insulin-induced kind, whereas rhoA P21-induced ruffling was apparently different from both the insulin- and HGF-induced kinds. Membrane ruffling was also induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), a protein kinase C-activating phorbol ester, but not by Ca2+ ionophore or microinjection of a dominant active Ki-ras P21 mutant (Ki-rasVal-12 P21). The phorbol ester-induced membrane ruffling was morphologically similar to the rhoA P21-induced kind and inhibited by microinjection of rho GDI or C3. These results indicate that rac P21 and rho GDI are involved in insulin-induced membrane ruffling and that rho P21 and rho GDI are involved in HGF- and phorbol ester-induced membrane rufflings.
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involvement of rho P21 small gtp binding protein and its regulator in the hgf induced cell motility
Oncogene, 1994Co-Authors: Kenji Takaishi, Takuya Sasaki, Masaki Kato, Wataru Yamochi, Shinya Kuroda, T Nakamura, Masatoshi Takeichi, Yoshimi TakaiAbstract:Hepatocyte growth factor (HGF) induced motility of cultured mouse keratinocytes (308R cells). This HGF-induced cell motility was inhibited by microinjection of either rho GDI, an inhibitory GDP/GTP exchange protein for rho P21 small GTP-binding protein, or a botulinum exoenzyme C3 which is known to selectively impair the function of rho P21 by ADP-ribosylating its effector domain. The rho GDI action was prevented by comicroinjection with the guanosine 5'-(3-0-thio)triphosphate (GTP gamma S)-bound active form of rhoA P21, and the C3 action was prevented by comicroinjection with a rhoA P21 mutant (rhoAIle41 P21) which is resistant to the C3 action. The HGF-induced cell motility was not inhibited by microinjection of a dominant negative rac1 P21 mutant (rac1Asn17 P21) or a dominant negative Ki-ras P21 mutant (Ki-rasAsn17 P21). Microinjection of the GTP gamma S-bound form of rac1 P21 or a dominant active Ki-ras P21 mutant (Ki-rasVal12 P21) did not induce cell motility. These results indicate that both rho P21 and rho GDI, but neither rac P21 nor ras P21, are involved in the HGF-induced cell motility. However, microinjection of the GTP gamma S-bound form of rhoA P21 alone did not induce cell motility in the absence of HGF, suggesting that activation of rho P21 is necessary but not sufficient for the HGF-induced cell motility. The HGF-induced cell motility was mimicked by 12-0-tetradecanoyl-phorbol-13-acetate, a protein kinase C-activating phorbol ester, but not by Ca2+ ionophore. The phorbol ester-induced cell motility was also inhibited by microinjection of rho GDI or C3. These results indicate that both rho P21 and rho GDI are also involved in the phorbol ester-induced cell motility.
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regulation of morphology by rho P21 and its inhibitory gdp gtp exchange protein rho gdi in swiss 3t3 cells
Journal of Biological Chemistry, 1993Co-Authors: Yasushi Miura, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, H Yaku, T Musha, Yoshimi TakaiAbstract:rho GDP dissociation inhibitor (GDI) is an inhibitory GDP/GTP exchange protein for a group of small GTP-binding proteins including at least rhoA P21, rhoB P21, rac1 P21, rac2 P21, and G25K. Microinjection of rho GDI into Swiss 3T3 cells made the cells round and refractile. This morphological change was accompanied by the disappearance of stress fibers. The rho GDI action was prevented by comicroinjection of rho GDI with the guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-bound form of rhoA P21, but not with the GTP gamma S-bound form of rhoA P21 lacking the C-terminal three amino acids, which was not post-translationally modified with lipids. The GTP gamma S-bound form of rac1 P21, the same form of G25K, the same form of smg P21B, or Ki-rasval12 P21 was ineffective. Microinjection of the bacterial ADP-ribosyltransferase C3 specific for rho P21 into Swiss 3T3 cells induced the similar changes of morphology and stress fibers. This C3 action was not prevented by comicroinjection of C3 with the GTP gamma S-bound form of rhoA P21, but was prevented by comicroinjection with the same form of a rhoA P21 mutant which was not ADP-ribosylated by C3. These results indicate that the rho GDI-rho P21 system regulates cell morphology presumably through the actomyosin system in Swiss 3T3 cells.
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involvement of rho P21 and its inhibitory gdp gtp exchange protein rho gdi in cell motility
Molecular and Cellular Biology, 1993Co-Authors: Kenji Takaishi, Akira Kikuchi, Takuya Sasaki, Shinya Kuroda, Ko Kotani, Yoshimi TakaiAbstract:Evidence is accumulating that rho P21, a ras P21-related small GTP-binding protein (G protein), regulates the actomyosin system. The actomyosin system is known to be essential for cell motility. In the present study, we examined the action of rho P21, its inhibitory GDP/GTP exchange protein (named rho GDI), its stimulatory GDP/GTP exchange protein (named smg GDS), and Clostridium botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho P21 and to impair its function, in cell motility (chemokinesis) of Swiss 3T3 cells. We quantitated the capacity of cell motility by measuring cell tracks by phagokinesis. Microinjection of the GTP gamma S-bound active form of rhoA P21 or smg GDS into Swiss 3T3 cells did not affect cell motility, but microinjection of rho GDI into the cells did inhibit cell motility. This rho GDI action was prevented by comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA P21 but not with the same form of rhoA P21 lacking the C-terminal three amino acids which was not posttranslationally modified with lipids. The rho GDI action was not prevented by Ki-rasVal-12 P21 or any of the GTP gamma S-bound form of other small GTP-binding proteins including rac1 P21, G25K, and smg P21B. Among these small G proteins, rhoA P21, rac1 P21, and G25K are known to be substrates for rho GDI. The rho GDI action was not prevented by comicroinjection of rho GDI with smg GDS. Microinjection of C3 into Swiss 3T3 cells also inhibited cell motility. These results indicate that the rho GDI-rho P21 system regulates cell motility, presumably through the actomyosin system.
Tsuyoshi Sakoda - One of the best experts on this subject based on the ideXlab platform.
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the functional domain of the stimulatory gdp gtp exchange protein smg gds which interacts with the c terminal geranylgeranylated region of rap1 krev 1 smg P21
Oncogene, 1992Co-Authors: K Kotani, Akira Kikuchi, K Doi, Shosei Kishida, Tsuyoshi Sakoda, K Kishi, Yoshimi TakaiAbstract:rap1/Krev-1/smg P21 (smg P21), a member of the small GTP-binding protein (G protein) superfamily, has a geranylgeranylated cysteine residue and clustered basic amino acids in the C-terminal region. The GDP/GTP exchange reaction of smg P21 is regulated by smg GDS, which is also active on Ki-ras P21 and rho P21. The C-terminal region of smg P21 is essential for its interaction with smg GDS. Moreover, smg P21 is phosphorylated by cyclic AMP- and cyclic GMP-dependent protein kinases at the serine residue between the polybasic region and the prenylated cysteine residue, and this phosphorylation initiates the smg GDS-induced smg P21 activation. Thus, the C-terminal cationic and hydrophobic region is important for the regulation of the smg P21 activity. In the present study, we attempted to determine the functional domain of smg GDS which interacts with the C-terminal region of smg P21 by use of a cross-link method and a site-directed mutagenesis method. The region of smg GDS cross-linked with the C-terminal region of smg P21B was residues 444-492, which is located at the C-terminal fifth of smg GDS. On deletion of these residues, smg GDS became inactive on smg P21B, Ki-ras P21 and rhoA P21. These results indicate that residues 444-492 of smg GDS are at least one of the domains which interact with the C-terminal region of its substrate small G proteins.
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a stimulatory gdp gtp exchange protein for smg P21 is active on the post translationally processed form of c ki ras P21 and rhoa P21
Proceedings of the National Academy of Sciences of the United States of America, 1991Co-Authors: T Mizuno, Tsuyoshi Sakoda, Kozo Kaibuchi, Takeshi Yamamoto, Motohiro Kawamura, Hiroyuki Fujioka, Yoshiharu Matsuura, Yoshimi TakaiAbstract:Abstract We have purified a stimulatory GDP/GTP exchange protein for smg P21A and -B, ras P21-like small GTP-binding proteins (G proteins), cloned its cDNA, and named it GDP dissociation stimulator (smg P21 GDS). We show here that smg P21 GDS is active not only on smg P21A and -B but also on c-Ki-ras P21 and rhoA P21, all of which are post-translationally processed. Furthermore, we show that smg P21 GDS is inactive on the post-translationally unprocessed form of these proteins and on the post-translationally unprocessed form of c-Ha-ras P21 and smg p25A. All of the small G proteins recognized by smg P21 GDS have a cDNA-predicted C-terminal "CAAX" motif (where C is cysteine, A is an aliphatic amino acid, and X is any amino acid) and a polybasic region upstream of this motif. These results suggest that smg P21 GDS is at least active on a group of small G proteins having these unique C-terminal structures. Moreover, they suggest that the C-terminal post-translational processing of these small G proteins, by farnesylation or geranylgeranylation of the C-terminal cysteine residue, removal of amino acids in positions denoted "AAX", and carboxyl methylation of the exposed cysteine residue, is important for the smg P21 GDS action.
