Molecular cloning of three malic acid related genes MdPEPC, MdVHA-A, MdcyME and their expression analysis in apple fruits

Malic acid (MA) in apple fruit is the predominant organic acid associated with taste, flavour and juice quality. In this study, three full-length cDNAs of MdPEPC, MdVHA-A and MdcyME were cloned from apple fruit. They encoded cytosolic phosphoenolpyruvate carboxylase (MdPEPC, EC 4.1.1.31), subunit A of vacuolar H+-ATPase (MdVHA, EC 3.6.1.3) and cytosolic NADP-dependent malic enzyme (MdcyME, EC 1.1.1.40), respectively, for MA synthesis, transportation and degradation. Real-time quantitative PCR discovered that the expression levels of three genes varied with development stages, and that their expression patterns differed between low acid (LA) and high acid (HA) genotypes. In addition, enzyme activity assay showed that PEPC and VHA contributed positively to MA accumulation during fruit development both in LA and HA, while cyME did negatively.