Several distinct genes encode nearly identical to 16 kDa proteolipids of the vacuolar H(+)-ATPase from Arabidopsis thaliana.
To understand the subcellular roles and the regulation of vacuolar H(+)-ATPases, we have begun to identify the genes encoding the major subunits and to determine their patterns of expression in Arabidopsis thaliana. Two distinct cDNAs (AVA-P1 and AVA-P2) and one genomic sequence (AVA-P3) encoding the 16 kDa subunit have been isolated. The 16 kDa proteolipid is a major component of the membrane integral sector that forms the proton conductance pathway and is required for assembly of the V-ATPase complex. Interestingly, the open reading frame of one full-length cDNA (AVA-P1) and a genomic sequence (AVA-P3) encoded an identical polypeptide of 164 amino acids with a molecular mass of 16,570. The deduced amino acid sequences of the two cDNAs were nearly identical (99%) and hydropathy plots suggested a molecule with four membrane-spanning domains characteristic of V-ATPase proteolipids. The three genes differed mainly in their codon usage and in their 3'-untranslated regions. The coding region of the genomic sequence, AVA-P3, was interrupted by two introns located at the codons for Cys-26 and Arg-121. The presence of additional 16 kDa proteolipid genes was suggested from several polymerase chain reaction (PCR)-amplified fragments that differed from one another in the size of the second intron. PCR 1 had an intron of ca. 800 bp and its identity as AVA-P4, a fourth member of the gene family, was confirmed from sequence analyses of an EST cDNA. The mRNAs of three genes (AVA-P1, AVA-P2 and AVA-P3) were detected in Arabidopsis leaf, root, flower and silique; yet expression of AVA-P1 and AVA-P2 was lower in roots. All three genes were expressed in light- or dark-grown seedlings; however mRNA levels of AVA-P2 were enhanced in etiolated plants. Arabidopsis thaliana, therefore, has at least four distinct genes encoding nearly identical 16 kDa proteolipids, and the enhanced expression of AVA-P2 transcript in etiolated seedlings suggests that an increase in V-ATPase could accompany cell expansion.
Perera
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Department of Plant Biology, University of Maryland, College Park 20742, USA.
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D000220:Adaptation, Biological; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics* / Q000528:radiation effects; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D015723:Gene Library; D017343:Genes, Plant; D018745:Genome, Plant; D008027:Light; D008969:Molecular Sequence Data; D005810:Multigene Family; D011487:Protein Conformation; D011510:Proteolipids / Q000235:genetics*; D006180:Proton-Translocating ATPases / Q000235:genetics*; D015183:Restriction Mapping; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D014018:Tissue Distribution; D014617:Vacuoles / Q000201:enzymology*
D018076:DNA, Complementary; D011510:Proteolipids; D006180:Proton-Translocating ATPases