Pan-genome-based characterization of SiGRAS transcription factors and prediction of hub genes correlated with plant height in foxtail millet (Setaria italica L.)

Abstract

The GRAS gene family, a class of plant-specific transcription factors, plays pivotal roles in diverse biological processes, including plant growth, development, and stress responses. Although the SiGRAS family has been previously characterized in foxtail millet, those studies were restricted to a single reference genome, overlooking critical genetic variations. To overcome the inherent limitations of gene family analysis using a single reference, we systematically investigated the SiGRAS gene family using pan-genome data generated from 110 foxtail millet accessions together with the xiaomi and Yugu1 genomes. Based on a rigorously filtered pan-genome catalog, we identified a total of 949 SiGRAS pan-genes, comprising 1 core, 20 near-core, 50 dispensable, and 878 private genes. Phylogenetic analysis classified these genes into 9 distinct subfamilies. Evolutionary analysis based on Ka/Ks ratios revealed that 8 SiGRAS genes underwent purifying selection, while SiGRAS17 and SiGRAS2 were subjected to positive selection in the majority of accessions. Notably, we observed that structural variations (SVs) significantly influenced the expression levels of SiGRAS63 , and altered both conserved domains and gene structure in specific accessions. Furthermore, we identified four hub genes correlated with plant height by integrating RNA-seq data with gene co-expression regulatory network. This study lays a foundation for understanding the diversity of the SiGRAS gene family and provides a valuable resource for future function studies of SiGRAS genes in foxtail millet.