Comparative analysis of mitochondrial citrate synthase gene structure, transcript level and enzymatic activity in acidless and acid-containing Citrus varieties

Most citrus (Citrus L. spp.) fruits accumulate a considerable amount of citric acid in the vacuoles of the juice sac cells. As part of research aimed to understand the mechanism of acid accumulation, we compared the gene structures and transcript levels of citrate synthase in sour lemon (high acid, C. limon (L.) Burm.), `Shamouti' orange (moderate acid, C. sinensis (L.) Osbeck) and sweet lime (acidless, C. limmetioides Tan.). Southern analyses suggested that a single gene for citrate synthase was present in the genomes of all three Citrus varieties. The gene structures seemed to be very similar, with minor differences in Shamouti orange. Overall, the transcript levels of citrate synthase were similar in sweet lime and sour lemon, and about 2-fold lower in Shamouti orange. The enzymatic activity of citrate synthase was compared between sour lemon and sweet lime. In sour lemon, the specific activity of the enzyme was induced early in fruit development and, in parallel with the increase in acid content, reached a maximal level, and did not diminish significantly towards fruit maturation; the pattern and level of activity detected during sweet lime fruit development were similar. These results suggest that the difference in acid accumulation between acidless and acid-containing fruits may not be attributed to changes in the activity of citrate synthase.