Pentatricopeptide repeat proteins: a socket set for organelle gene expression.

Cytoplasmic male sterility (CMS) and nucleus-controlled fertility restoration are widespread plant reproductive features that provide useful tools to exploit heterosis in crops. However, the molecular mechanism underlying this kind of cytoplasmic-nuclear interaction remains unclear. Here, we show in rice (Oryza sativa) with Boro II cytoplasm that an abnormal mitochondrial open reading frame, orf79, is cotranscribed with a duplicated atp6 (B-atp6) gene and encodes a cytotoxic peptide. Expression of orf79 in CMS lines and transgenic rice plants caused gametophytic male sterility. Immunoblot analysis showed that the ORF79 protein accumulates specifically in microspores. Two fertility restorer genes, Rf1a and Rf1b, were identified at the classical locus Rf-1 as members of a multigene cluster that encode pentatricopeptide repeat proteins. RF1A and RF1B are both targeted to mitochondria and can restore male fertility by blocking ORF79 production via endonucleolytic cleavage (RF1A) or degradation (RF1B) of dicistronic B-atp6/orf79 mRNA. In the presence of both restorers, RF1A was epistatic over RF1B in the mRNA processing. We have also shown that RF1A plays an additional role in promoting the editing of atp6 mRNAs, independent of its cleavage function.

Cytoplasmic Male Sterility of Rice with Boro II Cytoplasm Is Caused by a Cytotoxic Peptide and Is Restored by Two Related PPR Motif Genes via Distinct Modes of mRNA Silencing

MicroRNAs (miRNAs), a group of small non-coding RNAs, have recently become the subject of intense study. They are a class of post-transcriptional negative regulators playing vital roles in plant development and growth. However, little is known about their regulatory roles in the responses of trees to the stressful environments incurred over their long-term growth. Here, we report the cloning of small RNAs from abiotic stressed tissues of Populus trichocarpa (Ptc) and the identification of 68 putative miRNA sequences that can be classified into 27 families based on sequence homology. Among them, nine families are novel, increasing the number of the known Ptc-miRNA families from 33 to 42. A total of 346 targets was predicted for the cloned Ptc-miRNAs using penalty scores of </=2.5 for mismatched patterns in the miRNA:mRNA duplexes as the criterion. Six of the selected targets were validated experimentally. The expression of a majority of the novel miRNAs was altered in response to cold, heat, salt, dehydration, and mechanical stresses. Microarray analysis of known Ptc-miRNAs identified 19 additional cold stress-responsive Ptc-miRNAs from 14 miRNA gene families. Interestingly, we found that individual miRNAs of a family responded differentially to stress, which suggests that the members of a family may have different functions. These results reveal possible roles for miRNAs in the regulatory networks associated with the long-term growth of tree species and provide useful information for developing trees with a greater level of stress resistance.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-313X.2008.03497.x

Stress‐responsive microRNAs in Populus

Summary
RNA editing changes the sequence of many transcripts in plant organelles, but little is known about the molecular mechanisms determining the specificity of the process. In this study, we have characterized CLB19 (also known as PDE247), a gene that is required for editing of two distinct chloroplast transcripts, rpoA and clpP. Loss-of-function clb19 mutants present a yellow phenotype with impaired chloroplast development and early seedling lethality under greenhouse conditions. Transcript patterns are profoundly affected in the mutant plants, with a pattern entirely consistent with a defect in activity of the plastid-encoded RNA polymerase. CLB19 encodes a pentatricopeptide repeat protein similar to the editing specificity factors CRR4 and CRR21, but, unlike them, is implicated in editing of two target sites.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-313X.2008.03634.x

CLB19, a pentatricopeptide repeat protein required for editing of rpoA and clpP chloroplast transcripts.

In plants, RNA editing is a process for converting a specific nucleotide of RNA from C to U and less frequently from U to C in mitochondria and plastids. To specify the site of editing, the cis-element adjacent to the editing site functions as a binding site for the trans-acting factor. Genetic approaches using Arabidopsis thaliana have clarified that a member of the protein family with pentatricopeptide repeat (PPR) motifs is essential for RNA editing to generate a translational initiation codon of the chloroplast ndhD gene. The PPR motif is a highly degenerate unit of 35 amino acids and appears as tandem repeats in proteins that are involved in RNA maturation steps in mitochondria and plastids. The Arabidopsis genome encodes approximately 450 members of the PPR family, some of which possibly function as trans-acting factors binding the cis-elements of the RNA editing sites to facilitate access of an unidentified RNA editing enzyme. Based on this breakthrough in the research on plant RNA editing, I would like to discuss the possible steps of co-evolution of RNA editing events and PPR proteins.

RNA editing in plant organelles: machinery, physiological function and evolution

In this paper, we report a novel pentatricopeptide repeat (PPR) protein gene in rice. PPR, a characteristic repeat motif consisted of tandem 35 amino acids, has been found in various biological systems including plant. Sequence analysis revealed that the gene designated OsPPR1 consisted of an open reading frame of 2433 nucleotides encoding 810 amino acids that include 11 PPR motifs. Blast search result indicated that the gene did not align with any of the characterized PPR genes in plant. The OsPPR1 gene was found to contain a putative chloroplast transit peptide in the N-terminal region, suggesting that the gene product targets to the chloroplast. Southern blot hybridization indicated that the OsPPR1 is the member of a gene family within the rice genome. Expression analysis and immunoblot analysis suggested that the OsPPR1 was accumulated mainly in rice leaf. Antisense transgenic strategy was used to suppress the expression of OsPPR1 and the resulted transgenic rice showed the typical phenotypes of chlorophyll-deficient mutants; albinism and lethality. Cytological observation using microscopy revealed that the antisense transgenic plant contained a significant defect in the chloroplast development. Taken together, the results suggest that the OsPPR1 is a nuclear gene of rice, encoding the PPR protein that might play a role in the chloroplast biogenesis. This is the first report on the PPR protein required for the chloroplast biogenesis in rice.

Hongjoo Cho

Papers

OsPPR1, a pentatricopeptide repeat protein of rice is essential for the chloroplast biogenesis.