Abstract:

The caffeoyl-CoA O-methyltransferase （CCoAOMT） protein family belongs to the S-adenosyl-L-methionine （SAM）-dependent methyltransferase superfamily. Members of the CCoAOMT family are known to play key roles in plant lignin synthesis， as well as in growth and development. However， their function in Setaria italica has not been systematically studied. In this study， members of the SiCCoAOMT gene family were identified from S. italica genomic data， and bioinformatics analysis was conducted. In addition， the expression patterns of these genes under abiotic stress were determined. Five SiCCoAOMT genes in the SiCCoAOMT gene family were identified in the S. italica genome， and were located on chromosomes 2， 4， and 6. The physical and chemical properties differed among the five encoded putative proteins. In a phylogenetic analysis， the SiCCoAOMT genes of S. italica were divided into two subfamilies with homologous genes from Arabidopsis thaliana， Oryza sativa， and Setaria viridis. Members of the same subfamily shared high similarity in terms of conserved sequences and gene structures. All members of the CCoAOMT gene family contained the same conserved protein structure domains （motif 1， motif 2， motif 4， motif 5， and motif 6）， and members of the same subfamily had similar motifs. Prediction of cis-acting elements showed that SiCCoAOMT2 and SiCCoAOMT4 contained many elements responsive to light （Sp1）， plant hormones （ABRE， CGTCA-motif， and TGACG-motif） and abiotic stresses （ARE）. Collinearity analysis among S. italica， O. sativa， and S. viridis revealed that the CCoAOMT orthologs in S. italica and S. viridis have been subject to neutral selection and purifying selection， and the CCoAOMT orthologs in S. italica and O. sativa have been subject to positive selection. In a weighted gene co-expression network analysis， SiCCoAOMT2 and SiCCoAOMT4 were grouped in the yellowgreen module. To further verify the roles of SiCCoAOMT genes in stress responses， seedlings of the cultivar ‘Jingu 21’ were subjected to drought and low-temperature stresses， and the SiCCoAOMT transcript levels were analyzed by qRT-PCR. The results showed that the transcript levels of SiCCoAOMT2 and SiCCoAOMT4 were significantly up-regulated under both stress conditions. Based on single nucleotide polymorphism and insertion/deletion genotype information for 360 S. italica and 38 S.viridis germplasm resources， haplotype analysis showed that SiCCoAOMT5 was beneficial for the growth and development of S. italica grains， and contributed to the yield and stress resistance of S. italica. Overall， this study systematically characterized the SiCCoAOMT gene family in S. italica， identifying SiCCoAOMT2 and SiCCoAOMT4 as key genes involved in drought and cold stress responses. These findings provide a foundation for further exploration of the stress resistance and growth regulation mechanisms of S. italica.

Key words: Setaria italica, CCoAOMT gene family, abiotic stress, phylogenetic evolution, haplotype