Comparative transcriptome and co-expression network reveals key genes associated with mesocotyl elongation in foxtail millet (Setaria italica)

Abstract

Mesocotyl elongation characteristics decisively influence the emergence rate and deep-sowing tolerance of gramineous crops. However, there is limited research on the mechanism of mesocotyl elongation in foxtail millet under deep sowing conditions. To address this research gap, 119 foxtail millet accessions were used to analyze genetic diversity. Moreover the long mesocotyl variety Qinhuang 2 (Q) and the short mesocotyl variety Dungu 1 (D) were screened for pot experiments under different deep-sowing conditions (6, 8, and 10 cm) to investigate the relationship between seedling emergence ability and mesocotyl elongation. Mesocotyl tissues at a 10 cm sowing depth for two to six days were subjected to transcriptome sequencing and determination of endogenous hormone content. RNA-sequencing (RNA-Seq) was used to analyze the transcriptional mechanism of mesocotyl elongation under deep-sowing conditions, highlighting the key genes influencing several plant characteristics. The mesocotyl lengths of 119 foxtail millet materials exhibited a normal distribution and were divided into five groups, with the length decreasing from Group 1 to Group 5. Under deep-sowing conditions, the emergence rate of the long-mesocotyl variety Q was much higher than that of the short-mesocotyl variety D. The results indicate that under deep-sowing conditions, mesocotyl elongation is the main driving force for pushing the plumule through the soil surface. The endogenous IAA and ACC contents were significantly higher in the long mesocotyl variety Q than in the short mesocotyl variety D. When 10 µmol/L of IAA and 200 mg/L of ethephon were applied, the mesocotyls of both varieties were elongated. In contrast, the ABA content was the opposite, and no significant difference was observed in GA₃, indicating that IAA and ETH play a key role in the elongation of the mesocotyl of foxtail millet. In the co-expression network analysis, four related genes were identified in each of the “brown” and “purple” modules that were positively correlated with mesocotyl elongation. The expression levels of auxin-responsive proteins (IAA1, IAA17, and SAUR36) and ethylene genes EBF1 and EIL3 in Q were higher than those in D and increased with the elongation of the mesocotyl. Transcription factors MYC2, TGAL6, and PIF4 were involved in mesocotyl elongation. The “yellow” module was negatively correlated with mesocotyl elongation and contained transcription factors ERF, WRKY, and TGA2. Notably, by analyzing the levels of endogenous hormones and phenotypes, the expression patterns of the hub genes IAA17 and IAA1 have been found to be consistent with mesocotyl growth and show an overall upward trend. These findings provide insights into understanding the mechanism of mesocotyl elongation and highlight their potential application value in breeding foxtail millet to resist deep sowing.