Genome-wide identification, expression analysis of the MYC gene family in Stevia rebaudiana and the functional identification of SrMYC8 in SGs biosynthesis

Abstract

Stevia is an essential source of natural sweeteners-SGs. The MYC genes play crucial roles in modulating multiple biological processes. However, the functions of SrMYCs in Stevia remain poorly understood. SrMYCs were identified and characterized using bioinformatic methods for their physicochemical properties, phylogeny, genetic structures, conserved sequences, motif compositions, duplication patterns, chromosomal locations, and cis-acting elements. RNA-seq data were used to analyze the correlation between SrMYCs and SGs. Additionally, the expression patterns of SrMYCs were detected by RT-qPCR in various tissues and under diverse abiotic stresses. The function of SrMYC8 was validated through transient transformation, and the downstream target gene of SrMYC8 was investigated using Y1H experiments. 19 SrMYCs were identified from the Stevia genome and classified into 5 subgroups based on phylogeny. Cis-acting elements analysis revealed multiple elements involved in growth and development, light response, phytohormone signalling and abiotic stress, suggesting that SrMYCs play diverse physiological roles. Correlation analysis between SrMYCs and SGs identified SrMYC9/12/14/17 as potentially involved in SGs biosynthesis. The RT-qPCR analysis revealed that SrMYC8/9/12/14/17 differentially expressed in various tissueswith SrMYC8 was most highly expressed exhibiting the highest expression in mature leaves. Furthermore, these genes differentially expressed under various stresses, especially in response to MeJA and salts. Subcellular localization analysis confirmed SrMYC8 function within the nucleus. Transient transformation and Y1H experiments demonstrated that SrMYC8 enhances the expression of SrUGT91D2 by directly targeting its promoter. This study offers novel insights into the functions of the SrMYCs and regulation of SGs biosynthesis, providing valuable genetic resources for Stevia molecular breeding.