ABSTRACT

Pentatricopeptide repeat (PPR) proteins play an important role in post-transcriptional regulation of mitochondrial gene expression. Functions of many PPR proteins and their roles in plant growth and development remain unknown. Through characterization of an empty pericarp32 (emp32) mutant, we identified the function of Emp32 in mitochondrial intron splicing and seed development in maize. The loss-of-function mutant emp32 shows embryo lethality with severely arrested embryo and endosperm development, and over-expression of Emp32 rescues the embryo-lethality. EMP32 is a P-type PPR protein targeted to mitochondria. Loss of function in Emp32 dramatically decreases the splicing efficiency of nad7 intron 2, while complementation of Emp32 restores the splicing efficiency. Although nad7 intron 2 is partially spliced in the wild type, over-expression of Emp32 does not increase the splicing efficiency. The splicing deficiency of nad7 intron 2 blocks the assembly of mitochondrial complex I and dramatically reduces its activity, which may explain the embryo-lethality in emp32. In addition to the one copy of nad7 in the maize mitochondrial genome, we identified one to six copies of nad7 in the nuclear genomes in different maize inbred lines. These copies appear not to be expressed. Together, our results revealed that the P-type PPR protein EMP32 is required for the cis-splicing of nad7 intron 2 and seed development in maize.

摘要

五肽重复序列 (PPR) 蛋白在线粒体基因表达的转录后调控中起重要作用。许多 PPR 蛋白的功能及其在植物生长发育中的作用仍然未知。通过对空果皮32 ( emp32 ) 突变体的表征，我们确定了Emp32在玉米线粒体内含子剪接和种子发育中的功能。功能丧失的突变体emp32显示胚胎致死性，胚胎和胚乳发育严重停滞，Emp32 的过度表达挽救了胚胎致死性。EMP32 是一种靶向线粒体的 P 型 PPR 蛋白。Emp32 中的功能损失显着降低nad7内含子 2的剪接效率，而Emp32 的互补恢复了剪接效率。虽然nad7内含子2在野生型部分剪接的，过表达Emp32不增加剪接效率。nad7内含子 2的剪接缺陷阻止了线粒体复合物 I 的组装并显着降低了其活性，这可能解释了emp32 中的胚胎致死率。此外的一个副本nad7在玉米线粒体基因组，我们确定了一个以六份nad7在不同玉米自交系的核基因组中。这些副本似乎没有被表达。总之，我们的结果表明，玉米中nad7内含子 2的顺式剪接和种子发育需要 P 型 PPR 蛋白 EMP32 。