半胱氨酸脱硫酶(LCD)催化植物中信号分子硫化氢(H2S)的产生。在这项研究中,我们发现H2S可以抑制番茄(Solanumlycopersicum)果实成熟,而SlWRKY6在SlLCD1过表达的叶片中经历了差异的蛋白质过硫化。然后,进一步的研究表明,SlWRKY6可以在Cys396被H2S过硫化。通过构建slwrky6突变体和SlWRKY6-OE系,我们发现SlWRKY6通过激活成熟相关基因STAYGREEN1(SlSGR1)和衰老相关基因12(SlSAG12)的转录来正向调节叶片衰老和果实成熟。此外,SlWRKY6与激酶SlMAPK4相互作用并在Ser33处被磷酸化。双荧光素酶瞬时表达测定和电泳迁移率变化测定表明,SlWRKY6过硫化减弱了其对靶基因SlSGR1和SlSAG12的转录调节,而SlWRKY6通过SlMAPK4磷酸化激活了靶基因的转录以促进果实成熟。此外,我们提供了SlWRKY6过硫化作用减弱其SlMAPK4介导的磷酸化从而抑制番茄果实成熟的证据。通过瞬时表达SlWRKY6,SlWRKY6C396A,slwrky6水果中的slWRKY6S33A和slWRKY6S33D,我们发现SlWRKY6过硫化减弱了SlSGR1和SlSAG12的表达,从而延缓了番茄果实的成熟,而SlWRKY6磷酸化增加了靶基因的表达。随着番茄果实成熟,内源性H2S产量下降,而SlMAPK4表达增加。因此,我们的发现揭示了一个模型,在该模型中,由于未成熟果实中较高的内源性H2S水平而引起的SlWRKY6过硫化抑制了其激活SlSGR1和SlSAG12表达的能力,而SlWRKY6通过SlMAPK4磷酸化激活其转录活性,从而促进番茄果实成熟。
Cysteine desulfhydrase catalyses the generation of the signaling molecule hydrogen sulfide (H2S) in plants. In this study, we found that H2S can inhibit tomato (Solanum lycopersicum) fruit ripening and SlWRKY6 undergoes differential protein
persulfidation in SlLCD1-overexpressing leaves. Then, further study indicated that SlWRKY6 could be persulfidated by H2S at Cys396. By construction of slwrky6 mutants and SlWRKY6-OE lines, we found that SlWRKY6 positively regulates leaf senescence and fruit ripening by activating the transcription of ripening-related genes STAYGREEN 1 (SlSGR1) and Senescence-Associated Gene 12 (SlSAG12). In addition, SlWRKY6 interacted with kinase SlMAPK4 and was phosphorylated at Ser33. Dual-luciferase transient expression assays and electrophoretic mobility shift assays indicated that SlWRKY6
persulfidation attenuated its transcriptional regulation of target genes SlSGR1 and SlSAG12, whereas SlWRKY6 phosphorylation by SlMAPK4 activated the transcription of target genes to promote fruit ripening. Moreover, we provided evidence that SlWRKY6
persulfidation attenuated its SlMAPK4-mediated phosphorylation to inhibit tomato fruit ripening. By transient expression of SlWRKY6, SlWRKY6C396A, SlWRKY6S33A, and SlWRKY6S33D in slwrky6 fruits, we found that SlWRKY6
persulfidation attenuated the expression of SlSGR1 and SlSAG12 thereby delaying tomato fruit ripening, while SlWRKY6 phosphorylation increased the expression of target genes. As tomato fruits ripened, endogenous H2S production decreased, while SlMAPK4 expression increased. Therefore, our findings reveal a model in which SlWRKY6
persulfidation due to higher endogenous H2S levels in un-ripened fruit inhibits its ability to activate SlSGR1 and SlSAG12 expression, while SlWRKY6 phosphorylation by SlMAPK4 activates its transcriptional activity, thereby promoting tomato fruit ripening.