Premature chromatin condensation induced by loss of RCC1 is inhibited by GTP- and GTPgammaS-Ran, but not GDP-Ran.
RCC1 is a guanine nucleotide exchanging factor acting on nuclear G protein Ran. Premature chromatin condensation occurs in the temperature-sensitive rcc1- mutant of the BHK21 cell line, tsBN2, at the restrictive temperature. This observation can be explained if the premature activation of MPF is normally inhibited by GTP-Ran. In the absence of RCC1, GDP-Ran predominates, resulting in MPF activation. However, experiments with Ran mutants to determine whether GTP- or GDP-Ran prevents activation of MPF have yielded conflicting results. In order to clarify this point, we have microinjected nucleotide-bound Ran, instead of mutated Ran, into the nuclei of tsBN2 cells treated to reduce RCC1-mediated guanine nucleotide exchange. GTP-Ran, GTPgammaS-Ran, and GDP-Ran all inhibited chromatin condensation. However, the inhibition of chromatin condensation by GDP-Ran could be completely abolished by co-injection with GDP, but not GTP. Thus, we conclude that GTP-Ran blocks the activation of MPF and that hydrolysis of GTP is not required to prevent MPF activation.
Ohba
T
T
Department of Molecular Biology, Graduate School of Medical Science, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812, Japan.
Seki
T
T
Azuma
Y
Y
Nishimoto
T
T
D000818:Animals; D018797:Cell Cycle Proteins; D002460:Cell Line; D002843:Chromatin / Q000502:physiology*; D003001:Cloning, Molecular; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli; D019204:GTP-Binding Proteins / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D006153:Guanosine Diphosphate / Q000494:pharmacology*; D006160:Guanosine Triphosphate / Q000494:pharmacology*; D006801:Humans; D007668:Kidney; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature; D020931:ran GTP-Binding Protein
D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; C062521:RCC1 protein, human; D011994:Recombinant Proteins; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D020931:ran GTP-Binding Protein