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Proline and various betaines can function as osmoprotectants and cryoprotectants when accumulated in the cytoplasm of cells. Genetic engineering can raise levels of these compounds and thereby improve stress resistance;  Citrus  species are potential candidates for this. Before attempting such engineering, it is necessary to characterize the natural osmoprotectants of  Citrus  and related genera. We therefore surveyed 55 cultivated and wild species of the Aurantioideae, analyzing proline and betaines in leaves of mature trees. Some citrus relatives accumulated proline alone; others accumulated proline and proline betaine, as did all  Citrus  species studied. The levels of these two compounds ranged from about 20 to 100 μmol·g -1  dry mass, and were significantly inversely correlated. Proline betaine is known to be synthesized from proline and to be a better osmoprotectant. Because  Citrus  species all have more proline than proline betaine, there is scope for engineering more of the latter. Many species had small amounts of hydroxyproline betaine; other betaines were essentially absent. The lack of other betaines means that it would also be rational to engineer the accumulation of glycine betaine or similar compounds.

Proline Accumulation and Methylation to Proline Betaine in Citrus: Implications for Genetic Engineering of Stress Resistance