Genome-wide association analysis identifies a natural variation in basic helix-loop-helix transcription factor regulating ascorbate biosynthesis via D-mannose/L-galactose pathway in tomato

Abstract

Tomato ( Solanum lycopersicum ) is one of the highest-value vegetable crops worldwide. Understanding the genetic regulation of primary metabolite levels can inform efforts aimed toward improving the nutrition of commercial tomato cultivars, while maintaining key traits such as yield and stress tolerance. We identified 388 suggestive association loci (including 126 significant loci) for 92 metabolic traits including nutrition and flavor-related loci by genome-wide association study from 302 accessions in two different environments. Among them, an ascorbate quantitative trait locus TFA9 ( T OMATO F RUIT A SCORBATEON CHROMOSOME 9 ) co-localized with SlbHLH59 , which promotes high ascorbate accumulation by directly binding to the promoter of structural genes involved in the D-mannose/L-galactose pathway. The causal mutation of TFA9 is an 8-bp InDel, named InDel_8, located in the promoter region of SlbHLH59 and spanned a 5’UTR Py-rich stretch motif affecting its expression. Phylogenetic analysis revealed that differentially expressed SlbHLH59 alleles were selected during tomato domestication. Our results provide a dramatic illustration of how ascorbate biosynthesis can be regulated and was selected during the domestication of tomato. Furthermore, the findings provide novel genetic insights into natural variation of metabolites in tomato fruit, and will promote efficient utilization of metabolite traits in tomato improvement.

Perspectives of CRISPR/Cas-mediated cis-engineering in horticulture: unlocking the neglected potential for crop improvement

“…The mutation results in unbranched inflorescences with exceptionally large sepals 32 . An 8-bp insertion in the promoter of SlbHLH59 significantly increased its expression in accessions producing high ascorbic acid levels 37 . In apple (Malus × domestica), multiple repeats of a 23-bp motif in the promoter of MYB10 generate an autoregulatory locus, which is sufficient to account for increased expression and ectopic accumulation of anthocyanins in red-fleshed apples 38 .…”

Manipulation of Ascorbate Biosynthetic, Recycling, and Regulatory Pathways for Improved Abiotic Stress Tolerance in Plants

“…The tomato basic helix-loop-helix 59 (bHLH59) gene was recently identified as the likely candidate underlying the ascorbate quantitative trait locus TFA9 in tomato [110]. The bHLH59 transcription factor positively regulates the transcription of genes of the L-galactose pathway, including PMI, PMM, GMP1, GMP2, GMP3, GMP4, and GME1 [110]. Knock-down of the bHLH59 gene with RNAi in tomato reduced ascorbate to 65% of wild-type levels [110].…”

“…The bHLH59 transcription factor positively regulates the transcription of genes of the L-galactose pathway, including PMI, PMM, GMP1, GMP2, GMP3, GMP4, and GME1 [110]. Knock-down of the bHLH59 gene with RNAi in tomato reduced ascorbate to 65% of wild-type levels [110]. Increased expression of the bHLH59 gene in tomato increased ascorbate concentrations 1.5-fold and was associated with enhanced tolerance to methyl viologen-induced stress [110] (Table 3).…”

“…Knock-down of the bHLH59 gene with RNAi in tomato reduced ascorbate to 65% of wild-type levels [110]. Increased expression of the bHLH59 gene in tomato increased ascorbate concentrations 1.5-fold and was associated with enhanced tolerance to methyl viologen-induced stress [110] (Table 3).…”

Ascorbate and Thiamin: Metabolic Modulators in Plant Acclimation Responses

“…Recently, implementing genome wide association study (GWAS) on 302 tomato accessions identified a basic helix-loop-helix (bHLH) transcription factor, SlbHLH59, which positively regulates ascorbate content in tomato fruits [78]. The most similar protein to SlbHLH59 in Arabidopsis appears to be unfertilized embryo sac 12 (UNE12), regulating fertilization [78]. Further investigations are, however, needed to clarify whether this protein has links to the accumulation of ascorbate in Arabidopsis.…”