Cloning of vacuolar H+-ATPase subunit c genes from Japanese iris, and functional characterization in yeast

摘要

Five different isoforms (IrlVHA-cl-c5) of V-ATPase subunit c(VHA-c) were cloned from a Japanese iris (Iris lactea Pall. Var. chinensisFisch. Koidz) cDNA library using degenerate primers PCR and the5'-RACE technique. The sequence analysis showed the open readingframe (ORF) of the IrlVHA-c1-c5 to be 495 bp, corresponding to a pro-tein of 164 amino acids. Among the five isoforms, IrlVHA-c1 andIrlVHA-c2 are completely homologous. The IrlVHA-c protein is localizedat the vacuolar membrane as indicated by a green fluorescent protein(GFP) marker. Its over-expression in yeast could enhance yeast toleranceto NaCl stress. These results show that there are at least five genes en-coding different isoforms of IrlVHA-c in Japanese iris and IrlVHA-c isimportant for the function of V-ATPase.