OsHsp20-25, a small heat shock protein, from long-lived mRNA regulates seed size and germination rate in rice

The small heat shock protein OsHsp20-25, from long-lived mRNA, negatively regulates seed germination but positively regulates seed length. The mRNAs stored for a longer period in mature and dry seeds are called long-lived mRNA, which encodes diverse protein families and plays important roles in seed vigor. However, the specific contributions of individual mRNA populations to germination regulation remain poorly defined. In this study, we analyzed four small heat shock proteins, OsHsp20-17, OsHsp20-18, OsHsp20-20, and OsHsp20-25, from RNA sequencing data of rice seed. They were specifically expressed in the late stage of seed development and early stage of seed germination. Intriguingly, their transcripts retained detectable expression levels in one-year-old stored seeds. Four transgenic lines of OsHsp20s were generated, and only the OsHsp20-25 overexpression lines exhibited lower germination rates compared to the wild-type, accompanied by upregulated expression of OsABI5 and OsMFT2, and enhanced ABA sensitivity. Furthermore, OsHsp20-25-CRISPR mutants produced seeds of shorter length, but the overexpression line showed longer seeds. Biochemical and molecular evidence demonstrates that OsHsp20-25 interacted with OsHsp20-18, which also interacted with an F-box domain protein OsFBL45 in vivo and in vitro. Overall, OsHsp20, as a long-lived mRNA, plays important roles in seed development and seed germination.