H₂S mitigates chilling injury by downregulating PpbHLH113 to enhance phenylpropanoid biosynthesis in peach fruit

Abstract

Postharvest chilling injury (CI) severely limits peach fruit quality. Hydrogen sulfide (H₂S) has shown potential in mitigating CI, while its underlying transcriptional regulation mechanism remains insufficiently characterized. This study revealed that H₂S treatment effectively mitigated CI by promoting both the enzymatic activities and transcript levels of key phenylpropanoid biosynthesis enzymes (PAL, C4H, 4CL, and CHI), accompanied by increased accumulation of total phenolics and flavonoids. Conversely, the H₂S scavenger hypotaurine reversed these effects in peach fruit. Notably, PpbHLH113, a nuclear-localized newly identified bHLH family member, was upregulated by cold stress and downregulated by H₂S. Moreover, functional studies revealed that PpbHLH113 overexpression downregulated the expression of phenylpropanoid pathway genes. Further molecular interaction assays confirmed direct binding of PpbHLH113 to E-box cis-elements in the promoters of Pp4CL and PpCHI. Although deficient in transcriptional activation, PpbHLH113 functioned as a transcriptional repressor in dual-luciferase assays, significantly inhibiting the promoter activity of Pp4CL and PpCHI, thereby leading to the suppression of phenylpropanoid biosynthesis. Collectively, H₂S treatment protected peach fruit from chilling injury by downregulating PpbHLH113, thereby alleviating its transcriptional repression on key phenylpropanoid pathway genes, maintaining high levels of key enzyme activities and phenolic accumulation, and improving antioxidant capacity and membrane integrity.