The pentatricopeptide repeat (PPR) protein family is a large family characterized by tandem arrays of a degenerate 35-amino-acid motif whose members function as important regulators of organelle gene expression at the post-transcriptional level. Despite the roles of PPRs in RNA editing in organelles, their editing activities and the underlying mechanism remain obscure. Here, we show that a novel DYW motif-containing PPR protein, PPS1, is associated with five conserved RNA-editing sites of nad3 located in close proximity to each other in mitochondria, all of which involve conversion from proline to leucine in rice. Both pps1 RNAi and heterozygous plants are characterized by delayed development and partial pollen sterility at vegetative stages and reproductive stage. RNA electrophoresis mobility shift assays (REMSAs) and reciprocal competition assays using different versions of nad3 probes confirm that PPS1 can bind to cis-elements near the five affected sites, which is distinct from the existing mode of PPR-RNA binding because of the continuity of the editing sites. Loss of editing at nad3 in pps1 reduces the activity of several complexes in the mitochondrial electron transport chain and affects mitochondrial morphology. Taken together, our results indicate that PPS1 is required for specific editing sites in nad3 in rice.
A dual-localized PPR protein, OsPGL1, is required for simultaneous RNA editing in mitochondria and chloroplasts. OsPGL1 binds to two distinct target transcripts directly and cooperates with OsMORFs.
In land plants, the pentatricopeptide repeat (PPR) proteins form a large family involved in post-transcriptional processing of RNA in mitochondria and chloroplasts, which is critical for plant development and evolutionary adaption. Although studies showed a number of PPR proteins generally influence the editing of organellar genes, few of them were characterized in detail in rice. Here, we report a PLS-E subclass PPR protein in rice, PPR756, loss of function of which led to the abolishment of RNA editing events among three mitochondrial genes including atp6, ccmC, and nad7. Their defective C-to-U transformation then resulted in improper amino acid retention which could cause abortive pollen development. Furthermore, PPR756 could bind to the three target genes directly and interact with three OsMORFs (multiple organellar RNA editing factors): OsMORF1, OsMORF8-1, and OsMORF8-2. The knockout plants of PPR756 exhibited retarded growth and greener leaves during the early vegetative stages, along with sterile pollen and lower seed setting at the reproductive stage. These results established a role for PPR756 in rice development, participating in RNA editing of three various transcripts and cooperating with OsMORFs via an editosome manner in rice.
designed the experiments. Q.Z carried out most experiments. Y.X 22 performed the subcellular location and BiFC experiments. J.Huang contributed to the 23 protein purification and field management. K.Z performed the mitochondria isolation 24 and RNA Immunoprecipitation experiments. H.X and X.Q performed the data 25 analysis. L.Z contributed to the RNAi transgenic lines. Q.Z and J.Hu wrote the paper 26 with feedback from all authors. 27 28 One sentence summary: 39 PPR756 takes part in the RNA editing of three various mitochondrial transcripts, 40 cooperating with MORFs via editosome manner in rice. 41 42 ABSTRACT 43 In land plants, the pentatricopeptide repeat (PPR) proteins form a large family 44 accounting for post-transcriptional process in mitochondria and chloroplasts. 45 Although studies showed a number of PPR proteins generally influence the editing of 46 organellar genes acting as a member of editing complex, few of them were 47 characterized detailed in rice. Here, we reported a PLS-E subclass PPR protein in rice, 48 PPR756, loss function of which lead to the abolishment of RNA editing events among 49 three sites in mitochondrial genes including atp6 (ATP synthase F0 subunit 6), ccmC 50 (cytochrome c biogenesis C) and nad7 (NADH dehydrogenase subunit 7). The 51 defective C-to-U transformation of atp6-368, ccmC-236 and nad7-83 resulted in the 52 retention of proline, proline and serine codon respectively to the right leucine in rice. 53 Furthermore, PPR756 could bind to the three target genes directly and interacted with 54 three OsMORFs (multiple organellar RNA editing factors), OsMORF1, OsMORF8-1 55 and OsMORF8-2. The knock-out plants of PPR756 exhibited retarded growth and 56 greener leaves during the early stages, accompanied with sterile pollens at 57 reproductive stage. These results established a role for PPR756 in the RNA editing of 58 three various transcripts, cooperating with MORFs via editosome manner in rice.59 4
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