ISOLATION OF AN ADDITIONAL SOYBEAN CDNA-ENCODING YPT RAB-RELATED SMALL GTP-BINDING PROTEIN AND ITS FUNCTIONAL COMPARISON TO SYPT USING A YEAST YPT1-1 MUTANT/

We have previously reported the isolation of a gene from a soybean cDN A library encoding a Ypt/Rab-related small GTP-binding protein, Sypt. Here, we report the isolation of a second Ypt/Rab-related gene, design ated Srab2, from the same soybean cDNA library. And we compare the in vivo function of the two soybean genes utilizing a yeast ypt1-1 mutant . The Srab2 gene encodes 211 amino acid residues with a molecular mass of 23 169 Da. The deduced amino acid sequence of the Srab2 is closely related to the rat (76%) and human (75%) Rab2 proteins, but it shares relatively little homology to Sypt (46%) and Saccharomyces cerevisiae ypt proteins (41%). Genomic Southern blot analysis using the cDNA ins ert of Srab2 revealed that it belongs to a multigene family in the soy bean genome. The protein encoded by Srab2 gene, when expressed in Esch erichia coli, disclosed a GTP-binding activity. The expression pattern of the Srab2 gene is quite different from that of the Sypt gene. The Srab2 gene is predominantly expressed in the plumule region, while exp ression was very low in the other areas in soybean seedlings. On the o ther hand, the Sypt mRNA is not detectable in any tissues of soybean s eedlings grown in the dark. However, light significantly suppressed th e Srab2 gene expression, but enhanced the transcript levels of the Syp t gene in leaf and, at even higher levels, in root tissues. When the S rab2 and Sypt genes are introduced separately into a S cerevisiae defe ctive in vesicular transport function, the Srab2 gene cannot complemen t the temperature-sensitive yeast ypt1-1 mutation at all, in contrast to the Sypt gene. In conclusion, the difference of functional compleme ntation of the yeast mutation together with differential expression of the two genes suggest that the in vivo roles of the Srab2 and Sypt ge nes may be different in soybean cells.