The transcription factor CsbHLH60 relieves high-temperature inhibition of chlorophyll degradation in citrus.

Rising global temperature threatens fruit quality. As one of the most economically valuable fruits worldwide, citrus suffers from high-temperature-induced chlorophyll retention during ripening, significantly reducing commercial value. Comparative physiological analyses revealed that citrus fruit stored at 30°C exhibited markedly impaired chlorophyll degradation and failed to undergo normal degreening, in stark contrast to those maintained at 20°C. This temperature-dependent suppression of degreening was closely associated with a significant downregulation of key chlorophyll catabolic genes (CCGs). By integrating weighted gene co-expression network analysis (WGCNA) of a stay-green mutant, we identified the transcription factor CsbHLH60, a nuclear-localized transcriptional activator, from a chlorophyll-associated module. CsbHLH60 directly binds E-box motifs in the promoters of CsSGR and CsRCCR to drive their expression, a finding validated by ChIP-qPCR and EMSA assays. Transient overexpression of CsbHLH60 in citrus peel accelerated chlorophyll degradation and, importantly, overcame high-temperature-induced repression to restore degreening, whereas RNAi-mediated silencing at 20°C significantly impeded normal degreening of the peel. Mechanistically, high temperature imposes a dual "double-lock" suppression on CsbHLH60, concurrently repressing its transcription and promoting its protein degradation, thereby depleting functional activator levels. Collectively, our findings establish CsbHLH60 as a pivotal temperature-responsive regulator that directly couples temperature signaling to chlorophyll catabolism and is capable of overriding the high-temperature-induced suppression of chlorophyll degradation, thereby highlighting its potential as a molecular target for mitigating climate-driven declines in citrus fruit appearance quality.