RagA is a functional homologue of S. cerevisiae Gtr1p involved in the Ran/Gsp1-GTPase pathway.
Human RagA and RagB is reported to be 52% identical to a putative GTPase of Saccharomyces cerevisiae, Gtr1p. According to the reported nucleotide sequence, we amplified human RagA and RagBs cDNAs from the human B cell cDNA library with PCR. Both cDNAs rescued a cold sensitivity of S. cerevisiae, gtr1-11. Furthermore, we introduced into the cloned human RagA cDNA, the mutation 'T21L' corresponding to the gtr1-11 mutation which has been reported to suppress not only all of rcc1-, temperature-sensitive mutants of Ran/Gsp1p GTPase GDP/GTP-exchanging factor, but also rna1-1, a temperature-sensitive mutant of Ran/Gsp1p GTPase-activating protein. The resulting RagAgtr1-11 cDNA partially, but significantly, suppressed both rcc1- and rna1-1 mutations. These results indicated that RagA and RagBs are functional homologues of S. cervisiae Gtr1p. Interestingly, while wild-type human RagA and RagBs were localized within the cytoplasm, similar to S. cerevisiae Gtr1p, the mutated human RagAgtr1-11 corresponding to a dominant negative form of RagA was distributed in discrete speckles in the nucleus, being localized side by side with SC-35, a non-snRNP of the splicing complex. In contrast, a dominant positive form of RagA, Q66L was localized in the cytoplasm. Thus, RagA was suggested to shuttle between the cytoplasm and the nucleus, depending on the bound nucleotide state.
Hirose
E
E
Department of Molecular Biology, Graduate School of Medicine, Kyushu University, Fukuoka, Japan.
Nakashima
N
N
Sekiguchi
T
T
Nishimoto
T
T
D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins; D002467:Cell Nucleus / Q000737:chemistry / Q000201:enzymology; D003593:Cytoplasm / Q000737:chemistry / Q000201:enzymology; D004271:DNA, Fungal / Q000032:analysis; D004268:DNA-Binding Proteins / Q000235:genetics; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000032:analysis / Q000235:genetics*; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins; D016296:Mutagenesis / Q000502:physiology; D009687:Nuclear Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D020931:ran GTP-Binding Protein
D018797:Cell Cycle Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; C062521:RCC1 protein, human; C042027:RNA I; D012329:RNA, Bacterial; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C484342:RRAGA protein, human; C484345:RRAGB protein, human; D020559:Monomeric GTP-Binding Proteins; D020931:ran GTP-Binding Protein