Defects in secondary cell-wall synthesis mitigate the effects of lincomycin on early chloroplast development

Lincomycin (LIN)-mediated inhibition of protein synthesis in chloroplasts prevents greening of seedlings, represses the activity of photosynthesis-related genes in the nucleus including LHCB1.2, and induces the phenylpropanoid pathway, resulting in production of anthocyanins. In genomes uncoupled (gun) mutants, LHCB1.2 expression is maintained in the presence of LIN or other inhibitors of early chloroplast development. In a screen using concentrations of LIN lower than those employed to isolate gun mutants, we have identified happy on lincomycin (holi) mutants. Several holi mutants show increased tolerance to LIN, exhibiting de-repressed LHCB1.2 expression and chlorophyll synthesis in seedlings. The mutations responsible were identified by whole-genome SNP mapping, and most were found to affect the phenylpropanoid pathway. However, LHCB1.2 expression appears not to be directly regulated by phenylpropanoids, as indicated by metabolic profiling of mutants. The most potent holi mutant is defective in a subunit of cellulose synthase encoded by IRREGULAR XYLEM3, and comparative analysis of this and other cell-wall mutants establishes a link between secondary cell-wall integrity and tolerance to LIN, possibly involving altered ABA metabolism or sensing. Examination of differential gene expression in the Arabdidopsis thaliana irx3-2 mutant compared to wild type grown on MS or on MS supplemented with Lincomcycin

林可霉素（Lincomycin, LIN）介导的叶绿体蛋白质合成抑制作用，可抑制幼苗绿化、阻遏细胞核内LHCB1.2等光合相关基因的表达活性，并诱导苯丙烷途径，最终促成花青素生成。在基因组解偶联（genomes uncoupled, GUN）突变体中，即便存在LIN或其他叶绿体早期发育抑制剂，LHCB1.2的表达仍可维持。本研究采用低于分离GUN突变体所用浓度的LIN进行筛选，成功鉴定出林可霉素耐受（happy on lincomycin, HOLI）突变体。部分HOLI突变体对LIN的耐受性显著增强，其幼苗可解除LHCB1.2的表达抑制并恢复叶绿素合成。通过全基因组单核苷酸多态性（SNP）定位技术，研究人员鉴定出了对应的致病突变，其中多数突变会影响苯丙烷途径。但突变体的代谢组分析结果显示，LHCB1.2的表达似乎并非直接受苯丙烷类物质调控。活性最强的HOLI突变体存在纤维素合酶亚基编码缺陷，该亚基由不规则木质部3（IRREGULAR XYLEM3, IRX3）基因编码；对该突变体及其他细胞壁缺陷突变体的比较分析证实，次生细胞壁完整性与LIN耐受性之间存在关联，该关联可能涉及脱落酸（abscisic acid, ABA）代谢或信号感知的改变。本研究还对分别在MS培养基及添加林可霉素的MS培养基中培养的拟南芥（Arabidopsis thaliana）irx3-2突变体与野生型进行了差异基因表达分析。