Additional file 1 of Heat Stress Responsive Aux/IAA Protein, OsIAA29 Regulates Grain Filling Through OsARF17 Mediated Auxin Signaling Pathway
Creators: name: Chen, Zhanghao
name: Zhou, Wei
name: Guo, Xianyu
name: Ling, Sheng
name: Li, Wang
name: Wang, Xin
name: Yao, Jialing
Publisher: name: figshare
Year: 2024
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 1: Figure S1. Ambient temperature of growing areas during rice grain filling stage. 35 °C is marked as yellow dotted lines and 40 °C is marked as red dotted lines; TS, heat damage accumulated temperature during the whole filling stage; HS, heat damage hours during the whole filling stage; TS7, heat damage accumulated temperature during 0–7 DAP; HS7, heat damage hours during 0-7 DAP. Figure S2. Agronomic traits of OsIAA29-RNAi (Ri29-1, Ri29-2) under high temperature. (A) Relative expression level of OsIAA29-RNAi lines in ZH11. (B–C) 1000 grain weight and shrunken seed rate of WT and Ri-29 lines. (D) The WT and Ri-29 lines images of grains of mature seeds. Bar = 1cm. Figure S3. The results of yeast two-hybrid analysis of the interaction between OsIAA29 and OsARFs. The full-length OsIAA29 cDNA was cloned into a vector bearing the DNA binding domain (BD), and the full-length cDNA of OsARFs were cloned into a vector bearing an activation domain (AD). The transformants were grown on DDO (SD/-Leu/-Trp) and QDO (SD/-Leu/-Trp/-His/-Ade) plates. Figure S4. Identification of OsARF17 CRISPR line (osarf17) and overexpressed plant (OE17). (A) Mutation sites in osarf17-1 and osarf17-2, as compared with wild-type (WT) sequences, protospacer-adjacent motif sequences are shown in bold, and inserted or deleted nucleotides are indicated in red. (B) Relative expression level of overexpression materials lines in ZH11. (C) Detection of FLAG fusion protein in ZH11 and overexpression lines. Total proteins extracted from developing caryopses at 7-DAP were used for western blot analysis with an anti-FLAG antibody. (D–E) 1000 grain weight and shrunken seed rate of WT and osarf17 lines. Data are presented as means standard error (SE) of five biological replicates. P-values were calculated using two-tailed t-test. *P &lt; 0.05, **P &lt; 0.01. Figure S5. EMSA analysis OsARF17 bindings to the promoter of the target gene (OsPDIL1-1). Figure S6. OsIAA21 was a canonical AUX/IAA protein. (A) The structure of OsIAA21. The blue boxes represent the exons, the line represents the intron and the white boxes the UTRs. (B) Schematic diagram of the domains of OsIAA21. (C) Compared with other canonical AUX/IAA. Figure S7. Schematic diagram of the OsIAA29 response heat stress in rice endosperm. OsIAA29 plays upstream regulatory roles in endosperm development, and the accumulation of storage substances. When the seeds under heat stress conditions, the auxin level is low, the OsIAA29 protein is induced by heat stress, which leads to a high level of OsIAA29 protein, and OsARF17 are compete by OsIAA21 and OsIAA29 protein. When the seeds under normal conditions, the auxin level is normal, both OsIAA29 and OsIAA21 are degraded by a 26S proteasome-dependent mechanism, but OsIAA29 protein is slowly degraded, and the transcription activity of OsARF17 was released. Table S1. Primers for functional analysis of OsIAA29. Table S2. Detection of potential off-target sites for the sgRNAs. Table S3 Agronomic traits of osiaa29 under normal and high temperature in 2018. Table S4 Agronomic traits of WT and OsIAA29-RNAi lines under high temperature in 2019. Table S5 Agronomic traits of osarf17 under normal and high temperature in 2021. Table S6. qRT-PCR analysis of selected genes in osiaa29-1.