Alternative Splicing Underpins the ALMT9 Transporter Function for Vacuolar Malic Acid Accumulation in Apple

• Published in: Advanced science 2024-06, Vol.11 (22), p.e2310159-n/a
• Main Authors: Li, Chunlong, Krishnan, Srinivasan, Zhang, Mengxia, Hu, Dagang, Meng, Dong, Riedelsberger, Janin, Dougherty, Laura, Xu, Kenong, Piñeros, Miguel A., Cheng, Lailiang
• Format: Article
• Language: English
• Subjects: ALMT9; alternative splicing; Alternative Splicing - genetics; Amino acids; apple; Cell division; Fruit - genetics; Fruit - metabolism; fruit acidity; Fruits; Gene Expression Regulation, Plant - genetics; Genes; malate transport; Malates - metabolism; Malus - genetics; Malus - metabolism; Plant Proteins - genetics; Plant Proteins - metabolism; Proteins; Transcription factors; Vacuoles - genetics; Vacuoles - metabolism; Variance analysis
• ISSN: 2198-3844; 2198-3844
• DOI: 10.1002/advs.202310159

Vacuolar malic acid accumulation largely determines fruit acidity, a key trait for the taste and flavor of apple and other fleshy fruits. Aluminum‐activated malate transporter 9 (ALMT9/Ma1) underlies a major genetic locus, Ma, for fruit acidity in apple, but how the protein transports malate across the tonoplast is unclear. Here, it is shown that overexpression of the coding sequence of Ma1 (Ma1α) drastically decreases fruit acidity in “Royal Gala” apple, leading to uncovering alternative splicing underpins Ma1's function. Alternative splicing generates two isoforms: Ma1β is 68 amino acids shorter with much lower expression than the full‐length protein Ma1α. Ma1β does not transport malate itself but interacts with the functional Ma1α to form heterodimers, creating synergy with Ma1α for malate transport in a threshold manner (When Ma1β/Ma1α ≥ 1/8). Overexpression of Ma1α triggers feedback inhibition on the native Ma1 expression via transcription factor MYB73, decreasing the Ma1β level well below the threshold that leads to significant reductions in Ma1 function and malic acid accumulation in fruit. Overexpression of Ma1α and Ma1β or genomic Ma1 increases both isoforms proportionally and enhances fruit malic acid accumulation. These findings reveal an essential role of alternative splicing in ALMT9‐mediated malate transport underlying apple fruit acidity. Alternative splicing of aluminum‐activated malate transporter 9 (ALMT9/Ma1) generates two isoform proteins in apple: Ma1β being 68 amino acids shorter with much lower expression than the full‐length protein Ma1α. Ma1β does not transport malate itself, but interacts with the functional Ma1α to form heterodimers, creating synergy for vacuolar malate transport underlying apple fruit acidity.