OsNAC300 Positively Regulates Cadmium Stress Responses and Tolerance in Rice Roots

Abstract

Transcriptional regulation is involved in responding to cadmium (Cd) stress in plants. However, the molecular mechanisms of Cd stress responses regulated by transcription factors remain largely unknown in plants. In this study, a rice (Oryza sativa) NAC (no apical meristem [NAM]; Arabidopsis transcription activation factor [ATAF]; cup-shaped cotyledon [CUC]-related) family transcription factor, OsNAC300, was isolated and functionally characterized for its involvement in Cd stress responses and tolerance. OsNAC300 was localized to the nucleus. OsNAC300 was mainly expressed in roots and significantly induced by Cd treatment. Knockout of OsNAC300 resulted in increased sensitivity to Cd stress, while its overexpression lines enhanced tolerance to Cd stress. RNA-Seq analysis revealed that the mutant is impaired in regulating some important genes that were responsive to Cd stress in wild-type rice, such as the pathogenesis-related genes 10a (OsPR10a), OsPR10b, chalcone synthase 1 (OsCHS1), and several others, which was validated by reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis. Moreover, chromatin immunoprecipitation-qPCR assays and luciferase reporter assays demonstrated that OsNAC300 directly binds to the promoters of OsPR10a, OsPR10b, and OsCHS1 and activates their transcription. Overall, OsNAC300 is an important regulatory factor in Cd stress responses and tolerance in rice.

OsNAC15 Regulates Tolerance to Zinc Deficiency and Cadmium by Binding to OsZIP7 and OsZIP10 in Rice

“…Phylogenetic analysis showed that OsNAC15 is closely related to ONAC039, ONAC045, OsNAC300 and ONAC123, showing 49.05%, 39.06%, 36.93% and 25.56% identity with amino acids, respectively ( Figure S1A ). Among these TFs, OsNAC300 has been shown to be involved in CDT, and its expression is also induced by Zn deficiency [ 49 , 51 ]. We further conducted phylogenetic analysis in other species and showed that OsNAC15 is closely related to a hypothetical protein, OsJ_25622, sharing 93.36% identity ( Figure S1B ).…”

“…The knockout of OsNAC15 also greatly increased Cd translocation from root to shoot ( Figure 4 F), suggesting that OsNAC15 is an important regulator in Zn and Cd uptake and transport. Hu et al [ 49 ] reported that loss-of-function of OsNAC300 resulted in hypersensitivity to Cd stress, while the overexpression of OsNAC300 increased CDT. However, no difference was observed in the OsNAC300-OE lines and the wild-type in Cd uptake and transport.…”

“…TFs generally modulate downstream target genes by binding to specific motif elements in their promoters to perform specific physiological and biochemical functions. Although OsNAC15 and OsNAC300 are phylogenetically similar [ 49 ], they seem to have different DNA binding characteristics. OsNAC300 positively regulated the expression of OsCHS1 , OsPR10a and OsPR10b via binding their promoters, while OsNAC15 directly bound to the promoters of ZIP transporters including OsZIP7 and OsZIP10 and negatively regulated their transcriptions ( Figure 7 and Figure 8 ).…”

“…OsNAC300 positively regulated the expression of OsCHS1 , OsPR10a and OsPR10b via binding their promoters, while OsNAC15 directly bound to the promoters of ZIP transporters including OsZIP7 and OsZIP10 and negatively regulated their transcriptions ( Figure 7 and Figure 8 ). Hu et al [ 49 ] suggested that Cd uptake or transport-related transporters might not be the target genes of OsNAC300. Since they did not actually test the binding ability of OsNAC300 to any Cd transporters, direct binding of Cd transporters by OsNAC300 cannot be ruled out.…”

“…However, only OsNAC3 and OsNAC300 have been shown to regulate CDT through a transgenic approach. OsNAC300 is induced by Cd stress, mainly expressed in rice root [ 49 ]. The knockout of OsNAC300 decreases CDT, while its overexpression increases CDT.…”