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The P-class pentatricopeptide repeat protein binds to the 5’ untranslated and translated regions of chloroplast psbI-ycf12 transcript and is involved in mRNA stability

ABSTRACT: Chloroplast gene expression is controlled by numerous nuclear-encoded RNA-binding proteins. Among them, pentatricopeptide repeat (PPR) proteins are known to be a key player in posttranscriptional regulation in chloroplasts. However, the functions of many PPR proteins remain unknown. In this study, we characterized the function of a chloroplast-localized P-class PPR protein PpPPR_21 in Physcomitrella patens. Knockout (KO) mutants of PpPPR_21 exhibited a reduced growth of the protonemata and lower photosynthetic activity. Immuno-blot analysis and blue-native gel analysis showed a remarkable reduction of the photosystem II (PSII) reaction center protein and poorly formation of the PSII super-complexes in the KO mutants. To access whether PpPPR_21 is involved in the chloroplast gene expression, chloroplast genome-wide microarray analysis and northern blot hybridization were performed. These analyses indicated that the psbI-ycf12 transcript encoding the low molecular weight subunits of PSII, did not accumulate in the KO mutants while other psb transcripts accumulated at similar levels of WT and the KO mutants. A complemented PpPPR_21 KO moss transformed with the cognate full-length PpPPR_21 cDNA rescued the psbI transcript accumulation level. RNA binding experiments showed that the recombinant PpPPR_21 bound efficiently to the 5’-untraslated and translated region of the psbI mRNA. The present study suggests that PpPPR_21 may be essential for accumulation of a stable psbI-ycf12 mRNA.

ORGANISM(S): Physcomitrium patens

PROVIDER: GSE121554 | GEO | 2019/03/26

REPOSITORIES: GEO

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Two novel PLS-class pentatricopeptide repeat proteins are involved in the group II intron splicing of mitochondrial transcripts in the moss Physcomitrella patens

Project description:Pentatricopeptide repeat (PPR) proteins are RNA binding proteins that function in posttranscriptional regulation as gene-specific regulators of RNA metabolism in plant organelles. Plant PPR proteins are divided into four classes: P, PLS, E and DYW. The E- and DYW-class proteins are mainly implicated in RNA editing, whereas most of the P-class proteins predominantly participate in RNA cleavage, splicing and stabilization. In contrast, the functions of PLS-class proteins still remain obscure. Here, we report the function of PLS-class PpPPR_31 and PpPPR_9 in Physcomitrella patens. The knockout (KO) mutants of PpPPR_31 and PpPPR_9 exhibited slower protonema growth compared to the wild type. The PpPPR_31 KO mutants showed a considerable reduction in the splicing of nad5 intron 3 and atp9 intron 1. The PpPPR_9 KO mutants displayed severely reduced splicing of cox1 intron 3. An RNA electrophoresis mobility shift assay showed that the recombinant PpPPR_31 protein bound to the 5' region of nad5 exon 4 and the bulged-A region in domain VI of atp9 group II intron 1 while the recombinant PpPPR_9 bound to the translated region of ORF622 in cox1 intron 3. These results suggest that PLS-class PPR proteins may influence the splicing efficiency of mitochondrial group II introns.

2020-08-01 | GSE145050 | GEO

half_edit - A new predictive method for E+ PPR binding

Project description:ngs2018_04_half_edit-half_edit - Is there some transcriptomic defects in these different PPR KO mutants? - Identification of RNA editing defects in 3 differents KO mutants for E+ PPR. Results will be compared to different predictive methods in order to find out which one is the more accurate. Also looking for other transcriptomic defects in a pure-PPR.

2020-03-30 | GSE144674 | GEO

Genome-wide investigation of pentatricopeptide repeat (PPR) gene family in poplar and their expression analysis in response to biotic and abiotic stresses

Project description:Pentatricopeptide repeat (PPR) proteins, which are characterized by tandem 30-40 amino acid sequence motifs, constitute a large gene family in plants. These known PPR proteins have been identified to play important roles in organellar RNA metabolism and plant development in Arabidopsis and rice. However, functions of PPR genes in woody species remain still largely unknown. Here, we identified and characterized a total of 626 PPR genes containing PPR motifs in the poplar genome. A comprehensive genome-wide analysis of the poplar PPR gene family was performed, including chromosomal location, phylogenetic relationships, gene duplication. Transcriptomic analyses identified that 154 of the PtrPPR genes were induced by biotic and abiotic treatments, including Marssonina brunnea, salicylic acid (SA), methyl jasmonate (MeJA), wounding, cold and salinity. Quantitative RT-PCR analysis further confirmed the expression profiles of 11 PtrPPR genes under different stresses. Our results contribute to a more comprehensive understanding the roles of PPR proteins and provided an insight for improving the stress tolerance in poplar.

2018-01-31 | GSE109609 | GEO

The Arabidopsis NOT4A E3 ligase promotes PGR3 expression and regulates chloroplast translation

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.

2020-11-17 | GSE161571 | GEO

The Arabidopsis NOT4A E3 ligase promotes PGR3 expression and regulates chloroplast function

2021-09-09 | PXD022584 | Pride

Lysine acetylation of chloroplast proteins

Project description:In photosynthesis, light is absorbed by the thylakoid embedded light harvesting pigment protein complexes (LHCs) that surround the photosynthetic reaction centers, photosystem II (PSII) and photosystem I (PSI). The distribution of light energy between the photosystems is balanced through state transitions1,2, which refer to the phosphorylation-dependent association of the loosely bound (L) LHCII antenna trimer either to PSII or PSI 3,4. Apart from phosphorylation, other mechanisms regulating state transitions have not been reported this far. In this study we demonstrate that lysine (Lys) acetylation of chloroplast proteins is a prerequisite for state transitions in Arabidopsis thaliana. Knock-out mutants lacking a chloroplast acetyltransferase NSI (At1g32070; AtNSI, SNAT) show selective decreases in the Lys acetylation status of several photosynthetic proteins including PSI, PSII and LHCII subunits. Fluorescence measurements revealed that changes in the wavelength of illumination do not cause state transitions in the nsi mutants even though their LHCII phosphorylation status is not defected. Furthermore, biochemical analyses of thylakoid proteins and protein complexes showed that nsi plants are not able to accumulate the phosphorylation dependent PSI-LHCII megacomplex. Our results manifest that Lys acetylation by NSI has an integral role in the regulation of state transitions in Arabidopsis.

2018-07-03 | PXD007625 | Pride

Lysine acetylation of chloroplast proteins 2

2018-07-03 | PXD007630 | Pride

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

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.

2020-08-01 | GSE152579 | GEO

Transcriptional Responses of Karenia brevis to Phosphorus Depletion and Addition

Project description:The role of coastal nutrient sources in the persistence of Karenia brevis red tides in coastal waters of Florida is currently a contentious issue that warrants investigation into the nutrient physiology of this dinoflagellate species. The molecular mechanisms by which dinoflagellates respond to nutrient availability are essentially unexplored. The current study sought to determine if the transcriptome of K. brevis is responsive to phosphorus and informative of nutrient status. Stationary phase P-limited cultures did not exhibit up-regulation of transcripts considered to be hallmark of P depletion relative to nutrient-replete stationary phase cultures, despite rapid growth upon addition of the limiting nutrient. To study transcriptome responses to nutrient addition, the limiting nutrient was added to depleted cells and changes in global gene expression were assessed over the first 48 hours following nutrient addition. P-addition resulted in significant changes in approximately 4% of genes on the microarray, using a significance cutoff of 1.7-fold and p<10-4. By far, the earliest responding genes were dominated by pentatricopeptide repeat (PPR) proteins, which increased in expression up to 3-fold by 1 h following nutrient addition. PPR proteins are nuclear encoded proteins involved in chloroplast and mitochondria RNA processing. Correspondingly, other functions enriched in response to both nutrients were photosystem and ribosomal genes. Transcriptomic responses to the addition of P reflected primarily chloroplast functions. Even the earliest responding transcripts, those encoding PPR proteins, possess a 5’ trans-spliced leader sequence, suggesting that the observed rapid response of the transcriptome may be achieved, in part, through post-transcriptional mechanisms.

2011-08-09 | GSE28419 | GEO

Transcriptional Responses of Karenia brevis to Nitrogen Depletion and Addition

Project description:The role of coastal nutrient sources in the persistence of Karenia brevis red tides in coastal waters of Florida is currently a contentious issue that warrants investigation into the nutrient physiology of this dinoflagellate species. The molecular mechanisms by which dinoflagellates respond to nutrient availability are essentially unexplored. The current study sought to determine if the transcriptome of K. brevis is responsive to nitrogen and informative of nutrient status. Stationary phase N-limited cultures did not exhibit up-regulation of transcripts considered to be hallmark of N depletion relative to nutrient-replete stationary phase cultures, despite rapid growth upon addition of the limiting nutrient. To study transcriptome responses to nutrient addition, the limiting nutrient was added to depleted cells and changes in global gene expression were assessed over the first 48 hours following nutrient addition. N-addition resulted in significant changes in approximately 4% of genes on the microarray, using a significance cutoff of 1.7-fold and p<10-4. By far, the earliest responding genes were dominated by pentatricopeptide repeat (PPR) proteins, which increased in expression up to 3-fold by 4 h following nutrient addition. PPR proteins are nuclear encoded proteins involved in chloroplast and mitochondria RNA processing. Correspondingly, other functions enriched in response to both nutrients were photosystem and ribosomal genes. Transcriptomic responses to the addition of N reflected primarily chloroplast functions. Even the earliest responding transcripts, those encoding PPR proteins, possess a 5’ trans-spliced leader sequence, suggesting that the observed rapid response of the transcriptome may be achieved, in part, through post-transcriptional mechanisms.

2011-08-09 | GSE28362 | GEO

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