The AeMYBs responding to CMCS cooperatively regulate the phenylpropanoid pathway to participate in postharvest lignin...

Lignification is one of the key factors restricting the postharvest storage quality of okra. Analyzing the key factors inducing lignin deposition is beneficial for targeted regulation of the quality deterioration caused by postharvest lignification of okra. The aim of this study is to elucidate the molecular mechanisms of key v-myb avian myeloblastosis viral oncogene homolog (MYB) transcription factors involved in postharvest lignification of okra in response to carboxymethyl chitosan (CMCS). Subcellular localization experiments confirmed that AeMYB6, AeMYB315-1, and AeMYB35-2 are all nuclear localization proteins. The dual luciferase reporter gene (DLR) and yeast one hybrid (Y1H) experiments showed that the three AeMYBs can directly bind to promoters of multiple structural genes in the phenylpropanoid pathway, such as AePAL1/2/3/4 and AePOD1/2/3/4. Further transient overexpression (OE) experiments revealed that AeMYB6, AeMYB315-1, and AeMYB35-2 upregulated the expression of multiple genes (AePOD, Ae4CL, AeCAD, and AePAL) and enzyme activity in the phenylpropanoid pathway, including phenylalanine ammonia lyase (PAL), peroxidase (POD), 4-coumarin-CoA ligase (4CL), and cinnamoyl dehydrogenase (CAD). Metabolite analysis showed that lignin-related metabolites were reduced to varying degrees, such as coniferyl alcohol, sinapic acid, 4-hydroxycinnamic acid, caffeic acid, L-phenylalanine and ferulic acid. In summary, this study found that under the regulation of CMCS, AeMYB6, AeMYB315-1, and AeMYB35-2 cooperatively regulate the expression of multiple genes in the phenylpropanoid pathway, jointly regulating lignin accumulation. This result provides important scientific basis for revealing the mechanism of CMCS delaying postharvest lignification of okra.