The Phytocystatin AtCYS5 Enhances Seed Germination and Seedling Growth under Heat Stress Conditions.

Phytocystatins (PhyCYSs) are plant-specific proteinaceous inhibitors that are implicated in protein turnover and stress responses. Here, we characterized a PhyCYS from , which was designated AtCYS5. RT-qPCR analysis showed that the expression of in germinating seeds was induced by heat stress (HS) and exogenous abscisic acid (ABA) treatment. Analysis of the expression of the -glucuronidase reporter gene under the control of the promoter showed that expression during seed germination was induced by HS and ABA. Constitutive overexpression of driven by the cauliflower mosaic virus promoter led to enhanced HS tolerance in transgenic , which was characterized by higher fresh weight and root length compared to wild-type (WT) and knockout () plants grown under HS conditions. The HS tolerance of -overexpressing transgenic plants was associated with increased insensitivity to exogenous ABA during both seed germination and post-germination compared to WT and . Although no HS elements were identified in the 5'-flanking region of , canonical ABA-responsive elements (ABREs) were detected. was upregulated in ABA-treated protoplasts transiently co-expressing this gene and genes encoding bZIP ABRE-binding factors (ABFs and AREB3). In the absence of ABA, ABF1 and ABF3 directly bound to the ABREs in the promoter, which activated the transcription of this gene in the presence of ABA. These results suggest that an ABA-dependent pathway plays a positive role in the HS-responsive expression of during seed germination and post-germination growth.