Alternative splicing of ZmHsf23 modulates maize heat tolerance by regulating sHSPs and TIL1 expression

Heat stress is a major threat to maize (Zea mays L.) production worldwide. Heat shock transcription factors (HSFs) play vital roles in plant responses to heat stress. However, the molecular and genetic mechanisms underlying HSF-meditated thermotolerance in maize remain largely unexplored. In this study, we demonstrate that the alternative splicing of ZmHsf23 modulates heat stress tolerance in maize. Hsf23 produced two functional transcripts, Hsf23L and Hsf23S, which differ by the presence of a cryptic mini-exon in Hsf23L that is spliced out in Hsf23S. Both transcripts were strongly induced by heat stress. Mutants lacking Hsf23L alone (hsf23l) or both Hsf23L and Hsf23S (hsf23l23s) exhibited increased susceptibility to heat stress, whereas overexpression of Hsf23S enhanced heat stress tolerance in maize. Subsequently, we found that Hsf23S positively regulates heat stress tolerance by directly activating the transcription of three sHSP genes (Hsp16.9, Hsp17.2, and Hsp18a) and TIL1 gene. In addition, Hsf23L physically interacted with Hsf23S and enhanced the transcriptional activation of Hsf23S on the sHSPs and TIL1 promoters. Notably, genetic analysis suggested that co-overexpression of Hsf23L and Hsf23S further improves heat tolerance of the transgenic plants. Taken together, these results reveal two splicing variants of ZmHsf23 cooperatively regulate maize heat tolerance, thus highlighting potential value of ZmHsf23 in breeding heat-tolerant maize varieties.