A truncated ETHYLENE INSENSITIVE3-like protein, GhLYI, regulates senescence in cotton

Abstract Numerous endogenous and environmental signals regulate the intricate and highly orchestrated process of plant senescence. Ethylene (ET), which accumulates as senescence progresses, is a major promoter of leaf senescence. The master transcription activator ETHYLENE INSENSITIVE3 (EIN3) activates the expression of a wide range of downstream genes during leaf senescence. Here, we found that a unique EIN3-LIKE 1 (EIL1) gene, cotton LINT YIELD INCREASING (GhLYI), encodes a truncated EIN3 protein in upland cotton (Gossypium hirsutum L.) that functions as an ET signal response factor and a positive regulator of senescence. Ectopic expression or overexpression of GhLYI accelerated leaf senescence in both Arabidopsis (Arabidopsis thaliana) and cotton. Cleavage under targets and tagmentation (CUT&Tag) analyses revealed that SENESCENCE-ASSOCIATED GENE 20 (SAG20) was a target of GhLYI. Electrophoretic mobility shift assay (EMSA), yeast 1-hybrid (Y1H), and dual-luciferase transient expression assay confirmed that GhLYI directly bound the promoter of SAG20 to activate its expression. Transcriptome analysis revealed that transcript levels of a series of senescence-related genes, SAG12, NAC-LIKE, ACTIVATED by APETALA 3/PISTILLATA (NAP/ANAC029), and WRKY53, are substantially induced in GhLYI overexpression plants compared with wild-type (WT) plants. Virus-induced gene silencing (VIGS) preliminarily confirmed that knockdown of GhSAG20 delayed leaf senescence. Collectively, our findings provide a regulatory module involving GhLYI-GhSAG20 in controlling senescence in cotton.