Molecular Cloning of Tomato Plasma Membrane H-ATPase

Two cDNA clones (LHA1 and LHA2) from tomato (Lycopersicon esculentum) which likely encode isoforms of the plasma membrane H(+)-ATPase were isolated. The longest cDNA (3229 base pairs), LHA1, comprises an open reading frame that encodes a 956 amino acid, 105 kilodalton polypeptide with several potential transmembrane domains. In vitro transcription and translation of LHA1 yields a major translation product of approximately 100 kilodaltons that is immunoprecipitable with antiserum to the corn root plasma membrane H(+)-ATPase. LHA2 encodes a portion of a coding sequence that is 96% identical to LHA1, suggesting that LHA2 encodes an isoform of the H(+)-ATPase. Genomic DNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to a common set of six to eight restriction fragments at moderate stringency and to single distinct fragments at high stringency. LHA1 and LHA2 map to distinct sites on chromosomes three and six, respectively. RNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to 3.4 kilobase pair transcripts present in both leaves and roots, although the LHA2 transcript is relatively more abundant in leaves than in roots. These results indicate that in tomato as many as six to eight genes may encode the plasma membrane H(+)-ATPase, two of which are expressed at the level of mRNA in both roots and leaves.