Apple fruit acidity is genetically diversified by natural variations in three hierarchical epistatic genes: MdSAUR37, MdPP2CH and MdALMTII

Many efforts have been made to map quantitative trait loci (QTLs) to facilitate practical marker-assisted selection (MAS) in plants. In the present study, using MapQTL and BSA-seq (bulk segregant analysis using next generation sequencing) with two independent pedigree-based populations, we identified four major genome-wide QTLs responsible for apple fruit acidity. Candidate genes were screened in major QTL regions, and three functional gene markers, including a non-synonymous A/G single-nucleotide polymorphism (SNP) in the coding region of MdPP2CH, a 36-bp insertion in the promoter of MdSAUR37 and a previously reported SNP in MdALMTII, were validated to influence the malate content of apple fruits. In addition, MdPP2CH inactivated three vacuolar H⁺ -ATPases (MdVHA-A3, MdVHA-B2 and MdVHA-D2) and one aluminium-activated malate transporter (MdALMTII) via dephosphorylation and negatively influenced fruit malate accumulation. The dephosphotase activity of MdPP2CH was suppressed by MdSAUR37, which implied a higher hierarchy of genetic interaction. Therefore, the MdSAUR37/MdPP2CH/MdALMTII chain cascaded hierarchical epistatic genetic effects to precisely determine apple fruit malate content. An A/G SNP (-1010) on the MdMYB44 promoter region from a major QTL (qtl08.1) was closely associated with fruit malate content. The predicted phenotype values (PPVs) were estimated using the tentative genotype values of the gene markers, and the PPVs were significantly correlated with the observed phenotype values. Our findings provide an insight into plant genome-based selection in apples and will aid in conducting research to understand the fundamental physiological basis of quantitative genetics.