Characterization of the strictosidine synthase-like (SSL) gene family in foxtail millet reveals SiSSL4 confers...

Setaria italica (foxtail millet) is a climate-resilient cereal crop, yet the genetic components underlying its stress adaptability, particularly those in the strictosidine synthase-like (SSL) gene family, are poorly understood. To bridge this knowledge gap, we conducted the first genome-wide investigation of the SSL gene family in foxtail millet. We systematically identified 30 SiSSL genes, phylogenetically classified them into four distinct clades, and determined that the family evolved under strong purifying selection. Promoter and expression analyses revealed their regulatory complexity: SiSSL promoters are enriched with stress- and hormone-responsive cis-elements, and a subset of genes exhibited dynamic induction under drought, salt, cold, and hormone treatments. Focusing on the most promising candidate, SiSSL4, we demonstrated its dual localization to the nucleus and cell membrane and showed that heterologous overexpression of SiSSL4 in Arabidopsis confers robust drought tolerance. This enhanced tolerance is mechanistically linked to a potentiated antioxidant system, evidenced by significantly higher survival rates and elevated activities of key enzymes (POD, SOD, CAT). Our study not only provides a comprehensive genomic and functional framework for the SSL family in a stress-tolerant crop but also identifies and validates SiSSL4 as a key genetic resource for engineering drought-resistant cereals.