Identification and characterization of a natural SNP variant in ALTERNATIVE OXIDASE gene associated with cold stress tolerance in watermelon

Abstract

Alternative oxidase (AOX) is a mitochondrial enzyme encoded by a small nuclear gene family, which contains the two subfamilies, AOX1 and AOX2. In the present study on watermelon (Citrullus lanatus), only one ClAOX gene, belonging to AOX2 subfamily but having a similar gene structure to AtAOX1a, was found in the watermelon genome. The expression analysis suggested that ClAOX had the constitutive expression feature of AOX2 subfamily, but was cold inducible, which is normally considered an AOX1 subfamily feature. Moreover, one single nucleotide polymorphism (SNP) in ClAOX sequence, which led to the change from Lys (N) to Asn (K) in the 96th amino acids, was found among watermelon subspecies. Ectopic expression of two ClAOX alleles in the Arabidopsis aox1a knock-out mutant indicated that ClAOXK-expressing plants had stronger cold tolerance than aox1a mutant and ClAOXN-expressing plants. Our findings suggested watermelon genome contained a single ClAOX that possessed the expression features of both AOX1 and AOX2 subfamilies. A naturally existing SNP in ClAOX differentiated the cold tolerance of transgenic Arabidopsis plants, impling a possibility this gene might be a functional marker for stress-tolerance breeding.