Transcriptional profiles reveal the pivotal roles of RcHSFA2 and RcHSFA6b in mediating the heat stress response in Rosa chinensis

Heat stress significantly constrains rose ( Rosa spp.) growth and yield. Understanding the molecular mechanisms of heat stress response is essential for developing thermotolerant rose varieties. This study investigated the regulatory mechanisms underlying rose heat stress (42 °C) response by constructing gene regulatory networks (GRNs) based on temporal transcriptional patterns. Heat shock transcription factors (HSFs) showed significant enrichment across all heat treatment time points. Notably, RcHSFA2 and RcHSFA6b maintained the highest expression levels throughout and emerged as hub HSFs in GRNs. The overexpression of RcHSFA2 and RcHSFA6b in rose petals and tobacco reduced malondialdehyde and H 2 O 2 levels while enhancing antioxidant enzyme activities. RcHSFA2 and RcHSFA6b physically interact with RcHSFC1, and these RcHSFA–RcHSFC1 complexes contribute to the enhanced thermotolerance in rose petals compared with individual factors. In addition, RcHSFA6b activates RcHSFA2 expression, and both RcHSFA2 and RcHSFA6b activate the expression of the small heat shock protein RcHSP18.1a . RcHSFC1 functions as a co-activator, enhancing the activation of the RcHSP18.1a promoter by RcHSFA2 and RcHSFA6b. Importantly, RcHSP18.1a overexpression enhances thermotolerance in rose petals. This study establishes a cascading regulatory network for heat stress response mediated by RcHSFA2 and RcHSFA6b, providing a molecular framework for targeted genetic improvement of thermotolerance in rose.