Department of Plant Biology, University of Minnesota, St. Paul, Minnesota 55108, USA.
Genetics. 2011 Dec;189(4):1249-60. doi: 10.1534/genetics.111.133587. Epub 2011 Sep 21.
Mutations at the APM1 and APM2 loci in the green alga Chlamydomonas reinhardtii confer resistance to phosphorothioamidate and dinitroaniline herbicides. Genetic interactions between apm1 and apm2 mutations suggest an interaction between the gene products. We identified the APM1 and APM2 genes using a map-based cloning strategy. Genomic DNA fragments containing only the DNJ1 gene encoding a type I Hsp40 protein rescue apm1 mutant phenotypes, conferring sensitivity to the herbicides and rescuing a temperature-sensitive growth defect. Lesions at five apm1 alleles include missense mutations and nucleotide insertions and deletions that result in altered proteins or very low levels of gene expression. The HSP70A gene, encoding a cytosolic Hsp70 protein known to interact with Hsp40 proteins, maps near the APM2 locus. Missense mutations found in three apm2 alleles predict altered Hsp70 proteins. Genomic fragments containing the HSP70A gene rescue apm2 mutant phenotypes. The results suggest that a client of the Hsp70-Hsp40 chaperone complex may function to increase microtubule dynamics in Chlamydomonas cells. Failure of the chaperone system to recognize or fold the client protein(s) results in increased microtubule stability and resistance to the microtubule-destabilizing effect of the herbicides. The lack of redundancy of genes encoding cytosolic Hsp70 and Hsp40 type I proteins in Chlamydomonas makes it a uniquely valuable system for genetic analysis of the function of the Hsp70 chaperone complex.
在绿藻莱茵衣藻中 APM1 和 APM2 基因座的突变赋予对硫代磷酰胺和二硝基苯胺除草剂的抗性。apm1 和 apm2 突变之间的遗传相互作用表明基因产物之间存在相互作用。我们使用基于图谱的克隆策略鉴定了 APM1 和 APM2 基因。仅包含编码 I 型 Hsp40 蛋白的 DNJ1 基因的基因组 DNA 片段可拯救 apm1 突变体表型，使其对除草剂敏感并恢复温度敏感的生长缺陷。五个 apm1 等位基因的损伤包括错义突变和核苷酸插入和缺失，导致蛋白质改变或基因表达水平极低。编码已知与 Hsp40 蛋白相互作用的胞质 Hsp70 蛋白的 HSP70A 基因位于 APM2 基因座附近。在三个 apm2 等位基因中发现的错义突变预测了改变的 Hsp70 蛋白。包含 HSP70A 基因的基因组片段可拯救 apm2 突变体表型。结果表明，Hsp70-Hsp40 伴侣复合物的客户可能在增加衣藻细胞中的微管动力学方面发挥作用。伴侣系统无法识别或折叠客户蛋白会导致微管稳定性增加，并对除草剂破坏微管的作用产生抗性。编码胞质 Hsp70 和 Hsp40 I 型蛋白的基因在衣藻中没有冗余，这使得它成为研究 Hsp70 伴侣复合物功能的遗传分析的一个非常有价值的系统。
