The Chloroplast RNA Binding Protein CP31A Has a Preference for mRNAs Encoding the Subunits of the Chloroplast NDH Complex and is Required for Their Accumulation
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ABSTRACT:
PROVIDER: PRJNA639782 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR12023960.fastq.gz | Fastqsanger.gz | |||
| SRR12023961.fastq.gz | Fastqsanger.gz | |||
| SRR12023962.fastq.gz | Fastqsanger.gz | |||
| SRR12023963.fastq.gz | Fastqsanger.gz | |||
| SRR12023964.fastq.gz | Fastqsanger.gz |
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Project description:Chloroplast RNA processing requires a large number of nuclear-encoded RNA binding proteins (RBPs) that are imported post-translationally into the organelle. Most of these RBPs are highly specific for one or few target RNAs. By contrast, members of the chloroplast ribonucleoprotein family (cpRNPs) have a wider RNA target range. We here present a quantitative analysis of RNA targets of the cpRNP CP31A using digestion-optimized RNA co-immunoprecipitation with deep sequencing (DO-RIP-seq). This identifies the mRNAs coding for subunits of the chloroplast NDH complex as main targets for CP31A. We demonstrate using whole-genome gene expression analysis and targeted RNA gel blot hybridization that the ndh mRNAs are all down-regulated in cp31a mutants. This diminishes the activity of the NDH complex. Our findings demonstrate how a chloroplast RNA binding protein can combine functionally related RNAs into one post-transcriptional operon.
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Project description:Chloroplast function requires the coordinated action of nuclear- and chloroplast-derived proteins, including several hundred nuclear-encoded pentatricopeptide repeat (PPR) proteins that regulate plastid mRNA metabolism. Despite their large number and importance, regulatory mechanisms controlling PPR expression are poorly understood. Here we show that the Arabidopsis NOT4A ubiquitin-ligase positively regulates the expression of PROTON GRADIENT REGULATION 3 (PGR3), a PPR protein required for translating several thylakoid-localised photosynthetic components and ribosome subunits within chloroplasts. Loss of NOT4A function leads to a strong depletion of cytochrome b6f and NDH complexes, as well plastid 30S ribosomes, which reduces mRNA translation and negatively impacts photosynthetic capacity, causing pale-yellow and slow-growth phenotypes. Quantitative transcriptome and proteome analyses reveal that PGR3 is misregulated in not4a. We show that the molecular not4a defects mimic those of a pgr3 mutant, and that normal plastid function is restored through transgenic PGR3 expression. Our work identifies NOT4A as crucial for ensuring robust photosynthetic function during development and stress-response, through promoting PGR3 production and chloroplast translation.
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Project description:Chloroplast RNA processing requires a large number of nuclear-encoded RNA binding proteins (RBPs) that are imported post-translationally into the organelle. Most of these RBPs are highly specific for one or few target RNAs. By contrast, members of the chloroplast ribonucleoprotein family (cpRNPs) have a wider RNA target range. We here present a quantitative analysis of RNA targets of the cpRNP CP31A using digestion-optimized RNA co-immunoprecipitation with deep sequencing (DO-RIP-seq). This identifies the mRNAs coding for subunits of the chloroplast NAD(P)H dehydrogenase (NDH) complex as main targets for CP31A. We demonstrate using whole-genome gene expression analysis and targeted RNA gel blot hybridization that the ndh mRNAs are all down-regulated in cp31a mutants. This diminishes the activity of the NDH complex. Our findings demonstrate how a chloroplast RNA binding protein can combine functionally related RNAs into one post-transcriptional operon.
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