Pentatricopeptide repeat(PPR)proteins perform essential functions in post-transcriptional regulation of gene expression,particularly RNA editing and RNA splicing,in plant organelles.Although research on chloroplast bi...Pentatricopeptide repeat(PPR)proteins perform essential functions in post-transcriptional regulation of gene expression,particularly RNA editing and RNA splicing,in plant organelles.Although research on chloroplast biogenesis and development has been extensive,the functions of most PPR genes in this process in rice(Oryza sativa)remain incompletely understood.This study identifies a novel P-type PPR protein,YELLOW-GREEN LEAF AND SEEDLING LETHALITY(YGS),which localizes to rice chloroplasts.YGS shows predominant expression in leaves.The ygs mutants,generated through CRISPR/Cas9-mediated genome editing of the YGS gene,displayed yellow-green leaves and seedling lethality.These phenotypes corresponded with reduced pigment levels and disrupted chloroplast ultrastructure compared to wild-type plants.Furthermore,the expression of genes associated with chloroplast development and chlorophyll biosynthesis showed significant alterations in the ygs mutants.The absence of YGS function affected RNA editing of rpl2 and intron splicing of ycf3-1 in the plastid genome.Additionally,YGS demonstrated interaction with the chloroplast signal recognition particle protein Oscp SRP54b in yeast two-hybrid and bimolecular fluorescence complementation analyses.These results indicate that YGS participates in RNA editing and RNA splicing in chloroplasts,thus serving a vital role in rice chloroplast development.展开更多
Chloroplasts are essential for normal plant growth and development.In plants,pentatricopeptide repeat(PPR)proteins mediate RNA processing in chloroplasts.Here,we characterized a rice albino leaf 5(al5)mutant which exh...Chloroplasts are essential for normal plant growth and development.In plants,pentatricopeptide repeat(PPR)proteins mediate RNA processing in chloroplasts.Here,we characterized a rice albino leaf 5(al5)mutant which exhibits albinism during early leaf development.The MutMap+analysis and transformation experiments revealed that AL5 encodes a chloroplast-localized P-type PPR protein.The AL5 mutation resulted in the defective splicing of ribosomal protein L2(rpl2)and ribosomal protein S12(rps12),which are involved in the synthesis of chloroplast 50S and 30S ribosomal subunits,respectively.The RNA-electrophoretic mobility shift assay(REMSA)further demonstrated that AL5 directly binds to rpl2 transcripts.Finally,disruption of AL5 led to reduced expression of plastid-encoded polymerase(PEP)-dependent plastid genes and nuclear-encoded photosynthetic genes.Notably,the albino al5 mutant phenotype was regulated by low temperature.These results suggest that AL5 participates in plastid RNA splicing and plays an important role in chloroplast development in rice.展开更多
Photosynthesis occurs mainly in chloroplasts,whose development is regulated by proteins encoded by nuclear genes.Among them,pentapeptide repeat(PPR)proteins participate in organelle RNA editing.Although there are more...Photosynthesis occurs mainly in chloroplasts,whose development is regulated by proteins encoded by nuclear genes.Among them,pentapeptide repeat(PPR)proteins participate in organelle RNA editing.Although there are more than 450 members of the PPR protein family in rice,only a few affect RNA editing in rice chloroplasts.Gene editing technology has created new rice germplasm and mutants,which could be used for rice breeding and gene function study.This study evaluated the functions of OsPPR9 in chloroplast RNA editing in rice.The osppr9 mutants were obtained by CRISPR/Cas9,which showed yellowing leaves and a lethal phenotype,with suppressed expression of genes associated with chloroplast development and accumulation of photosynthetic-related proteins.In addition,loss of OsPPR9 protein function reduces the editing efficiency of rps8-C182,rpoC2-C4106,rps14-C80,and ndhB-C611 RNA editing sites,which affects chloroplast growth and development in rice.Our data showed that OsPPR9 is highly expressed in rice leaves and encodes a DYW-PPR protein localized in chloroplasts.Besides,the OsPPR9 protein was shown to interact with OsMORF2 and OsMORF9.Together,our findings provide insights into the role of the PPR protein in regulating chloroplast development in rice.展开更多
Pentatricopeptide repeat(PPR)proteins are involved in nearly all aspects of post-transcriptional process-ing in plant mitochondria and plastids,playing vital roles in plant growth,development,cytoplasmic male sterilit...Pentatricopeptide repeat(PPR)proteins are involved in nearly all aspects of post-transcriptional process-ing in plant mitochondria and plastids,playing vital roles in plant growth,development,cytoplasmic male sterility restoration,and responses to biotic and abiotic stresses.Over the last three decades,significant advances have been made in understanding the functions of PPR proteins and the primary mechanisms through which they mediate post-transcriptional processing.This review aims to summarize these ad-vancements,highlighting the mechanisms by which PPR proteins facilitate RNA editing,intron splicing,and RNA maturation in the context of organellar gene expression.We also present the latest progress in PPR engineering and discuss its potential as a biotechnological tool.Additionally,we discuss key chal-lenges and questions that remain in PPR research.展开更多
Gamete maturation is critical for fertility of both animals and plants;however,the molecular mechanisms underlying these processes remain poorly understood in plants.Here,we report the Female Gamete Maturation 1(FGM1/...Gamete maturation is critical for fertility of both animals and plants;however,the molecular mechanisms underlying these processes remain poorly understood in plants.Here,we report the Female Gamete Maturation 1(FGM1/rPPR4),a component of mitoribosome large subunit in Arabidopsis directly interacts with the mitochondrial protein GAMETE CELL DEFECTIVE 1(GCD1)and plays an essential role in female gamete maturation and subsequent zygote–embryo transition and endosperm development.We reveal that FGM1/rPPR4,assisted by GCD1,is an essential factor for female gamete maturation.We also confirm that female gamete maturation is necessary for the capacity of post-fertilization zygote–embryo transition and endosperm development,but not for double fertilization,indicating that essential mechanisms are established during female gamete maturation to provide a molecular basis for seed development initiation and plant fertility.展开更多
Cotton is one of the most important textile fibers worldwide.As crucial agronomic traits,leaves play an essential role in the growth,disease resistance,fiber quality,and yield of cotton plants.Pentatricopeptide repeat...Cotton is one of the most important textile fibers worldwide.As crucial agronomic traits,leaves play an essential role in the growth,disease resistance,fiber quality,and yield of cotton plants.Pentatricopeptide repeat(PPR)proteins are a large family of nuclear-encoded proteins involved in organellar or nuclear RNA metabolism.Using a virus-induced gene silencing assay,we found that cotton plants displayed variegated yellow leaf phenotypes with decreased chlorophyll content when expression of the PPR gene GhCTSF1 was silenced.GhCTSF1 encodes a chloroplast-localized protein that contains only two PPR motifs.Disruption of GhCTSF1 substantially reduces the splicing efficiency of rpoC1 intron 1 and ycf3 intron 2.Loss of function of the GhCTSF1 ortholog EMB1417 causes splicing defects in rpoC1 and ycf3-2,leading to impaired chloroplast structure and decreased photosynthetic rates in Arabidopsis.We also found that GhCTSF1 interacts with two splicing factors,GhCRS2 and GhWTF1.Defects in GhCRS2 and GhWTF1 severely affect intron splicing of rpoC1 and ycf3-2 in cotton,leading to defects in chloroplast development and a reduction in photosynthesis.Our results suggest that GhCTSF1 is specifically required for splicing rpoC1 and ycf3-2 in cooperation with GhCRS2 and GhWTF1.展开更多
Pentatricopeptide repeat(PPR) proteins function in post-transcriptional regulation of organellar gene expression. Although several PPR proteins are known to function in chloroplast development in rice(Oryza sativa), t...Pentatricopeptide repeat(PPR) proteins function in post-transcriptional regulation of organellar gene expression. Although several PPR proteins are known to function in chloroplast development in rice(Oryza sativa), the detailed molecular functions of many PPR proteins remain unclear.Here, we characterized a rice young leaf white stripe(ylws) mutant, which has defective chloroplast development during early seedling growth.Map-based cloning revealed that YLWS encodes a novel P-type chloroplast-targeted PPR protein with 11 PPR motifs. Further expression analyses showed that many nuclear-and plastid-encoded genes in the ylws mutant were significantly changed at the RNA and protein levels. The ylws mutant was impaired in chloroplast ribosome biogenesis and chloroplast development under low-temperature conditions. The ylws mutation causes defects in the splicing of atpF, ndhA, rpl2,and rps12, and editing of ndhA, ndhB, and rps14transcripts. YLWS directly binds to specific sites in the atpF, ndhA, and rpl2 pre-mRNAs. Our results suggest that YLWS participates in chloroplast RNA group II intron splicing and plays an important role in chloroplast development during early leaf development.展开更多
RNA editing is a posttranscriptional process that is important in mitochondria and plastids of higher plants. All RNA editing-specific trans-factors reported so far belong to PLS-class of pentatricopeptide repeat(PPR)...RNA editing is a posttranscriptional process that is important in mitochondria and plastids of higher plants. All RNA editing-specific trans-factors reported so far belong to PLS-class of pentatricopeptide repeat(PPR)proteins. Here, we report the map-based cloning and molecular characterization of a defective kernel mutant dek39 in maize. Loss of Dek39 function leads to delayed embryogenesis and endosperm development, reduced kernel size, and seedling lethality. Dek39 encodes an E subclass PPR protein that targets to both mitochondria and chloroplasts, and is involved in RNA editing in mitochondrial NADH dehydrogenase3(nad3) at nad3-247 and nad3-275. C-to-U editing of nad3-275 is not conserved and even lost in Arabidopsis, consistent with the idea that no close DEK39 homologs are present in Arabidopsis. However, the amino acids generated by editing nad3-247 and nad3-275 are highly conserved in many other plant species, and the reductions of editing at these two sites decrease the activity of mitochondria NADH dehydrogenase complex I,indicating that the alteration of amino acid sequence is necessary for Nad3 function. Our results indicate that Dek39 encodes an E sub-class PPR protein that is involved in RNA editing of multiple sites and is necessary for seed development of maize.展开更多
As an important yield component,rice tiller number controls panicle number and determines grain yield.Regulation of rice tiller number by chloroplast pentatricopeptide repeat(PPR)proteins has not been reported previou...As an important yield component,rice tiller number controls panicle number and determines grain yield.Regulation of rice tiller number by chloroplast pentatricopeptide repeat(PPR)proteins has not been reported previously.Here,we report the rice reduced culm number22(rcn22)mutant,which produces few tillers owing to suppressed tiller bud elongation.Map-based cloning revealed that RCN22 encodes a chloroplast-localized P-type PPR protein.We found that RCN22 specifically binds to the 5′UTR of RbcL mRNA(encoding the large subunit of Rubisco)and enhances its stability.The reduced abundance of RbcL mRNA in rcn22 leads to a lower photosynthetic rate and decreased sugar levels.Consequently,transcript levels of DWARF3(D3)and TEOSINTE BRANCHED1(TB1)(which encode negative regulators of tiller bud elongation)are increased,whereas protein levels of the positive regulator DWARF53(D53)are decreased.Furthermore,high concentrations of sucrose can rescue the tiller bud growth defect of the rcn22 mutant.On the other hand,TB1 directly binds to the RCN22 promoter and downregulates its expression.The tb1/rcn22 double mutant shows a tillering phenotype similar to that of rcn22.Our results suggest that the TB1–RCN22–RbcL module plays a vital role in rice tiller bud elongation by affecting sugar levels.展开更多
Most of the reported P-type pentatricopeptide repeat(PPR) proteins play roles in organelle RNA stabilization and splicing. However, P-type PPRs involved in both RNA splicing and editing have rarely been reported, and ...Most of the reported P-type pentatricopeptide repeat(PPR) proteins play roles in organelle RNA stabilization and splicing. However, P-type PPRs involved in both RNA splicing and editing have rarely been reported, and their underlying mechanism remains largely unknown. Here, we report a rice floury endosperm22(flo22) mutant with delayed amyloplast development in endosperm cells. Map-based cloning and complementation tests demonstrated that FLO22 encodes a mitochondrion-localized P-type PPR protein.Mutation of FLO22 resulting in defective transsplicing of mitochondrial nad1 intron 1 and perhaps causing instability of mature transcripts affected assembly and activity of complex Ⅰ, and mitochondrial morphology and function. RNA-seq analysis showed that expression levels of many genes involved in starch and sucrose metabolism were significantly down-regulated in the flo22mutant compared with the wild type, whereas genes related to oxidative phosphorylation and the tricarboxylic acid cycle were significantly upregulated. In addition to involvement in splicing as a P-type PPR protein, we found that FLO22 interacted with DYW3, a DYW-type PPR protein, and they may function synergistically in mitochondrial RNA editing. The present work indicated that FLO22 plays an important role in endosperm development and plant growth by participating in nad1 maturation and multi-site editing of mitochondrial messager RNA.展开更多
Pentatricopeptide repeat （PPR） proteins comprise a large family in higher plants and modulate organellar gene expression by participating in various aspects of organellar RNA metabolism. In rice, the family contains...Pentatricopeptide repeat （PPR） proteins comprise a large family in higher plants and modulate organellar gene expression by participating in various aspects of organellar RNA metabolism. In rice, the family contains 477 members, and the majority of their functions remain unclear. In this study, we isolated and characterized a rice mutant, white stripe leaf （wsl）, which displays chlorotic striations early in development. Map-based cloning revealed that WSL encodes a newly identified rice PPR protein which targets the chloroplasts. In wsl mutants, PEP-dependent plastid gene expression was significantly down-regulated, and plastid rRNAs and translation products accumulate to very low levels. Consistently with the observations, wsl shows a strong defect in the splicing of chloroplast transcript rpl2, resulting in aberrant transcript accumulation and its product reduction in the mutant. The wsl shows enhanced sensitivity to ABA, salinity, and sugar, and it accumulates more H2O2 than wild-type. These results suggest the reduced translation efficiency may affect the response of the mutant to abiotic stress.展开更多
RNA editing sites in plant mitochondria and plastids are addressed by pentatricopeptide repeat （PPR） proteins with E or E and DYW domains, which recognize a specific nucleotide motif upstream of the edited nucleotid...RNA editing sites in plant mitochondria and plastids are addressed by pentatricopeptide repeat （PPR） proteins with E or E and DYW domains, which recognize a specific nucleotide motif upstream of the edited nucleotide. In addition, some sites require MORF proteins for efficient RNA editing. Here, we assign the novel E domain-containing PPR protein, MEF13, as being required for editing at eight sites in Arabidopsis thaliana. A SNP in ecotype C24 altering the editing level at only one of the eight target sites was located by genomic mapping. An EMS mutant allele of the gene for MEF13 was identified in a SNaPshot screen of a mutated plant population. At all eight target sites of MEF13, editing levels are reduced in both morf3 and morf8 mutants, but at only one site in morfl mutants, suggesting that specific MEF13-MORF interactions are required. Yeast two-hybrid analyses detect solid connections of MEF13 with MORF1 and weak contact with MORF3 proteins. Yeast three-hybrid （Y3H） analysis shows that the presence of MORF8 enhances the connection between MEF13 and MORF3, suggesting that a MORF3-MORF8 heteromer may form stably or transiently to establish interaction with MEF13.展开更多
Pentatricopeptide repeat(PPR)proteins play important roles in the post-transcriptional modification of organellar RNAs in plants.However,the function of most PPR proteins remains unknown.Here,we characterized the rice...Pentatricopeptide repeat(PPR)proteins play important roles in the post-transcriptional modification of organellar RNAs in plants.However,the function of most PPR proteins remains unknown.Here,we characterized the rice(Oryza sativa L.)chlorophyll deficient 4(cde4)mutant which exhibits an albino phenotype during early leaf development,with decreased chlorophyll contents and abnormal chloroplasts at low-temperature(20℃).Positional cloning revealed that CDE4 encodes a P-type PPR protein localized in chloroplasts.In the cde4 mutant,plastid-encoded polymerase(PEP)-dependent transcript levels were significantly reduced,but transcript levels of nuclear-encoded genes were increased compared to wild-type plants at 20℃.CDE4 directly binds to the transcripts of the chloroplast genes rpl2,ndhA,and ndhB.Intron splicing of these transcripts was defective in the cde4 mutant at 20℃,but was normal at 32℃.Moreover,CDE4 interacts with the guanylate kinase VIRESCENT 2(V2);overexpression of V2 enhanced CDE4 protein stability,thereby rescuing the cde4 phenotype at 20℃.Our results suggest that CDE4 participates in plastid RNA splicing and plays an important role in rice chloroplast development under lowtemperature conditions.展开更多
基金supported by the National Natural Science Foundation of China(32201784,32072048,and U2004204)the Natural Science Foundation of Shandong Province,China(ZR2020QC111 and ZR2022QC176)the Talent Introduction Project of Dezhou University,China(2020xjrc207)。
文摘Pentatricopeptide repeat(PPR)proteins perform essential functions in post-transcriptional regulation of gene expression,particularly RNA editing and RNA splicing,in plant organelles.Although research on chloroplast biogenesis and development has been extensive,the functions of most PPR genes in this process in rice(Oryza sativa)remain incompletely understood.This study identifies a novel P-type PPR protein,YELLOW-GREEN LEAF AND SEEDLING LETHALITY(YGS),which localizes to rice chloroplasts.YGS shows predominant expression in leaves.The ygs mutants,generated through CRISPR/Cas9-mediated genome editing of the YGS gene,displayed yellow-green leaves and seedling lethality.These phenotypes corresponded with reduced pigment levels and disrupted chloroplast ultrastructure compared to wild-type plants.Furthermore,the expression of genes associated with chloroplast development and chlorophyll biosynthesis showed significant alterations in the ygs mutants.The absence of YGS function affected RNA editing of rpl2 and intron splicing of ycf3-1 in the plastid genome.Additionally,YGS demonstrated interaction with the chloroplast signal recognition particle protein Oscp SRP54b in yeast two-hybrid and bimolecular fluorescence complementation analyses.These results indicate that YGS participates in RNA editing and RNA splicing in chloroplasts,thus serving a vital role in rice chloroplast development.
基金supported by the Open Competition Program of Top Ten Critical Priorities of Agricultural Science and Technology Innovation for the 14th Five-Year Plan of Guangdong Province(2022SDZG05)the Guangdong Province Rural Revitalization Strategy Special Fund Seed Industry Revitalization Project(2022-NJS-15-001)the Key-Area Research and Development Program of Guangdong Province(2022B0202060005).
文摘Chloroplasts are essential for normal plant growth and development.In plants,pentatricopeptide repeat(PPR)proteins mediate RNA processing in chloroplasts.Here,we characterized a rice albino leaf 5(al5)mutant which exhibits albinism during early leaf development.The MutMap+analysis and transformation experiments revealed that AL5 encodes a chloroplast-localized P-type PPR protein.The AL5 mutation resulted in the defective splicing of ribosomal protein L2(rpl2)and ribosomal protein S12(rps12),which are involved in the synthesis of chloroplast 50S and 30S ribosomal subunits,respectively.The RNA-electrophoretic mobility shift assay(REMSA)further demonstrated that AL5 directly binds to rpl2 transcripts.Finally,disruption of AL5 led to reduced expression of plastid-encoded polymerase(PEP)-dependent plastid genes and nuclear-encoded photosynthetic genes.Notably,the albino al5 mutant phenotype was regulated by low temperature.These results suggest that AL5 participates in plastid RNA splicing and plays an important role in chloroplast development in rice.
基金funded by the Central Public-Interest Scientific Institution Basal Research Fund,China(CPSIBRF-CNRRI-202111 and CPSIBRF-CNRRI-202110)the Agricultural Science and Technology Innovation Program,Chinese Academy of Agricultural Sciences(ASTIP)+1 种基金the Project of State Key Laboratory of Rice Biology,China(2020ZZKT10205)the Key Research and Development Project of China Rice Research Institute(CNRRI-2020-01)。
文摘Photosynthesis occurs mainly in chloroplasts,whose development is regulated by proteins encoded by nuclear genes.Among them,pentapeptide repeat(PPR)proteins participate in organelle RNA editing.Although there are more than 450 members of the PPR protein family in rice,only a few affect RNA editing in rice chloroplasts.Gene editing technology has created new rice germplasm and mutants,which could be used for rice breeding and gene function study.This study evaluated the functions of OsPPR9 in chloroplast RNA editing in rice.The osppr9 mutants were obtained by CRISPR/Cas9,which showed yellowing leaves and a lethal phenotype,with suppressed expression of genes associated with chloroplast development and accumulation of photosynthetic-related proteins.In addition,loss of OsPPR9 protein function reduces the editing efficiency of rps8-C182,rpoC2-C4106,rps14-C80,and ndhB-C611 RNA editing sites,which affects chloroplast growth and development in rice.Our data showed that OsPPR9 is highly expressed in rice leaves and encodes a DYW-PPR protein localized in chloroplasts.Besides,the OsPPR9 protein was shown to interact with OsMORF2 and OsMORF9.Together,our findings provide insights into the role of the PPR protein in regulating chloroplast development in rice.
基金supported by the National Natural Science Foundation of China(project no.32101640 and 32230075).
文摘Pentatricopeptide repeat(PPR)proteins are involved in nearly all aspects of post-transcriptional process-ing in plant mitochondria and plastids,playing vital roles in plant growth,development,cytoplasmic male sterility restoration,and responses to biotic and abiotic stresses.Over the last three decades,significant advances have been made in understanding the functions of PPR proteins and the primary mechanisms through which they mediate post-transcriptional processing.This review aims to summarize these ad-vancements,highlighting the mechanisms by which PPR proteins facilitate RNA editing,intron splicing,and RNA maturation in the context of organellar gene expression.We also present the latest progress in PPR engineering and discuss its potential as a biotechnological tool.Additionally,we discuss key chal-lenges and questions that remain in PPR research.
基金supported by the National Natural Science Foundation of China(32130031,32270360,31991201,and 31800265)the Natural Science Foundation of Hubei Province(2024AFA010,2024AFE007).
文摘Gamete maturation is critical for fertility of both animals and plants;however,the molecular mechanisms underlying these processes remain poorly understood in plants.Here,we report the Female Gamete Maturation 1(FGM1/rPPR4),a component of mitoribosome large subunit in Arabidopsis directly interacts with the mitochondrial protein GAMETE CELL DEFECTIVE 1(GCD1)and plays an essential role in female gamete maturation and subsequent zygote–embryo transition and endosperm development.We reveal that FGM1/rPPR4,assisted by GCD1,is an essential factor for female gamete maturation.We also confirm that female gamete maturation is necessary for the capacity of post-fertilization zygote–embryo transition and endosperm development,but not for double fertilization,indicating that essential mechanisms are established during female gamete maturation to provide a molecular basis for seed development initiation and plant fertility.
基金supported by the National Natural Science Foundation of China(32170367 and 32000146)the Shaanxi Fundamental Science Research Project for Chemistry&Biology(22JHZ007)+2 种基金the Department of Science and Technology Innovation Team Project of Shaanxi Provincial(2024RS-CXTD-72)the Fundamental Research Funds for the Central Universities(2020TS053)the Excellent Graduate Training Program of Shaanxi Normal University(LHRCCX23185).
文摘Cotton is one of the most important textile fibers worldwide.As crucial agronomic traits,leaves play an essential role in the growth,disease resistance,fiber quality,and yield of cotton plants.Pentatricopeptide repeat(PPR)proteins are a large family of nuclear-encoded proteins involved in organellar or nuclear RNA metabolism.Using a virus-induced gene silencing assay,we found that cotton plants displayed variegated yellow leaf phenotypes with decreased chlorophyll content when expression of the PPR gene GhCTSF1 was silenced.GhCTSF1 encodes a chloroplast-localized protein that contains only two PPR motifs.Disruption of GhCTSF1 substantially reduces the splicing efficiency of rpoC1 intron 1 and ycf3 intron 2.Loss of function of the GhCTSF1 ortholog EMB1417 causes splicing defects in rpoC1 and ycf3-2,leading to impaired chloroplast structure and decreased photosynthetic rates in Arabidopsis.We also found that GhCTSF1 interacts with two splicing factors,GhCRS2 and GhWTF1.Defects in GhCRS2 and GhWTF1 severely affect intron splicing of rpoC1 and ycf3-2 in cotton,leading to defects in chloroplast development and a reduction in photosynthesis.Our results suggest that GhCTSF1 is specifically required for splicing rpoC1 and ycf3-2 in cooperation with GhCRS2 and GhWTF1.
基金supported by the Key Laboratory of Biology, GeneticsBreeding of Japonica Rice in the Mid-lower Yangtze River, Ministry of Agriculture, China+5 种基金Jiangsu Plant Gene Engineering Research CenterJiangsu Collaborative Innovation Center for Modern Crop ProductionThis research was also supported by grants from the National Natural Science Foundation (92035301)Jiangsu Science and Technology Development Program (BE2021360)Jiangsu Agricultural Science and Technology Innovation Fund Project (SCX (19)1079)the Fundamental Research Funds for the Central Universities (JCQY201902)。
文摘Pentatricopeptide repeat(PPR) proteins function in post-transcriptional regulation of organellar gene expression. Although several PPR proteins are known to function in chloroplast development in rice(Oryza sativa), the detailed molecular functions of many PPR proteins remain unclear.Here, we characterized a rice young leaf white stripe(ylws) mutant, which has defective chloroplast development during early seedling growth.Map-based cloning revealed that YLWS encodes a novel P-type chloroplast-targeted PPR protein with 11 PPR motifs. Further expression analyses showed that many nuclear-and plastid-encoded genes in the ylws mutant were significantly changed at the RNA and protein levels. The ylws mutant was impaired in chloroplast ribosome biogenesis and chloroplast development under low-temperature conditions. The ylws mutation causes defects in the splicing of atpF, ndhA, rpl2,and rps12, and editing of ndhA, ndhB, and rps14transcripts. YLWS directly binds to specific sites in the atpF, ndhA, and rpl2 pre-mRNAs. Our results suggest that YLWS participates in chloroplast RNA group II intron splicing and plays an important role in chloroplast development during early leaf development.
基金supported by the National Natural Science Foundation of China (91435206 31421005)+1 种基金National Key Technologies Research & Development ProgramSeven Major Crops Breeding Project (2016YFD0101803, 2016YFD0100404)the 948 project (2016-X33)
文摘RNA editing is a posttranscriptional process that is important in mitochondria and plastids of higher plants. All RNA editing-specific trans-factors reported so far belong to PLS-class of pentatricopeptide repeat(PPR)proteins. Here, we report the map-based cloning and molecular characterization of a defective kernel mutant dek39 in maize. Loss of Dek39 function leads to delayed embryogenesis and endosperm development, reduced kernel size, and seedling lethality. Dek39 encodes an E subclass PPR protein that targets to both mitochondria and chloroplasts, and is involved in RNA editing in mitochondrial NADH dehydrogenase3(nad3) at nad3-247 and nad3-275. C-to-U editing of nad3-275 is not conserved and even lost in Arabidopsis, consistent with the idea that no close DEK39 homologs are present in Arabidopsis. However, the amino acids generated by editing nad3-247 and nad3-275 are highly conserved in many other plant species, and the reductions of editing at these two sites decrease the activity of mitochondria NADH dehydrogenase complex I,indicating that the alteration of amino acid sequence is necessary for Nad3 function. Our results indicate that Dek39 encodes an E sub-class PPR protein that is involved in RNA editing of multiple sites and is necessary for seed development of maize.
基金Biological Breeding-National Science and Technology Major Project of China(2023ZD0406801)Innovation Program of Chinese Academy of Agricultural Sciences(CAAS-CSCB-202402)+1 种基金National Natural Science Foundation of China(32270373,32270653)Agricultural Variety Improvement Project of Shandong Province,China(2021LZGC020).
文摘As an important yield component,rice tiller number controls panicle number and determines grain yield.Regulation of rice tiller number by chloroplast pentatricopeptide repeat(PPR)proteins has not been reported previously.Here,we report the rice reduced culm number22(rcn22)mutant,which produces few tillers owing to suppressed tiller bud elongation.Map-based cloning revealed that RCN22 encodes a chloroplast-localized P-type PPR protein.We found that RCN22 specifically binds to the 5′UTR of RbcL mRNA(encoding the large subunit of Rubisco)and enhances its stability.The reduced abundance of RbcL mRNA in rcn22 leads to a lower photosynthetic rate and decreased sugar levels.Consequently,transcript levels of DWARF3(D3)and TEOSINTE BRANCHED1(TB1)(which encode negative regulators of tiller bud elongation)are increased,whereas protein levels of the positive regulator DWARF53(D53)are decreased.Furthermore,high concentrations of sucrose can rescue the tiller bud growth defect of the rcn22 mutant.On the other hand,TB1 directly binds to the RCN22 promoter and downregulates its expression.The tb1/rcn22 double mutant shows a tillering phenotype similar to that of rcn22.Our results suggest that the TB1–RCN22–RbcL module plays a vital role in rice tiller bud elongation by affecting sugar levels.
基金supported by grants from the National Key R&D Program of China (2021YFF1000200)National Natural Science Foundation of China (31901513)+6 种基金the “JBGS” Project of Seed Industry Revitalization in Jiangsu Province (JBGS [2021]008)Jiangsu Province Agriculture Independent Innovation Fund Project (CX(19)1002)the Fundamental Research Funds for the Central Universities (KJQN202005)the China Postdoctoral Science Foundation (2019M661864)also supported by the Key Laboratory of Biology, Genetics, and Breeding of Japonica Rice in Mid-lower Yangtze River, Ministry of Agriculture, Chinathe Jiangsu Collaborative Innovation Center for Modern Crop ProductionNational Observation and Research Station of Rice Germplasm Resources, Nanjing, Jiangsu。
文摘Most of the reported P-type pentatricopeptide repeat(PPR) proteins play roles in organelle RNA stabilization and splicing. However, P-type PPRs involved in both RNA splicing and editing have rarely been reported, and their underlying mechanism remains largely unknown. Here, we report a rice floury endosperm22(flo22) mutant with delayed amyloplast development in endosperm cells. Map-based cloning and complementation tests demonstrated that FLO22 encodes a mitochondrion-localized P-type PPR protein.Mutation of FLO22 resulting in defective transsplicing of mitochondrial nad1 intron 1 and perhaps causing instability of mature transcripts affected assembly and activity of complex Ⅰ, and mitochondrial morphology and function. RNA-seq analysis showed that expression levels of many genes involved in starch and sucrose metabolism were significantly down-regulated in the flo22mutant compared with the wild type, whereas genes related to oxidative phosphorylation and the tricarboxylic acid cycle were significantly upregulated. In addition to involvement in splicing as a P-type PPR protein, we found that FLO22 interacted with DYW3, a DYW-type PPR protein, and they may function synergistically in mitochondrial RNA editing. The present work indicated that FLO22 plays an important role in endosperm development and plant growth by participating in nad1 maturation and multi-site editing of mitochondrial messager RNA.
文摘Pentatricopeptide repeat （PPR） proteins comprise a large family in higher plants and modulate organellar gene expression by participating in various aspects of organellar RNA metabolism. In rice, the family contains 477 members, and the majority of their functions remain unclear. In this study, we isolated and characterized a rice mutant, white stripe leaf （wsl）, which displays chlorotic striations early in development. Map-based cloning revealed that WSL encodes a newly identified rice PPR protein which targets the chloroplasts. In wsl mutants, PEP-dependent plastid gene expression was significantly down-regulated, and plastid rRNAs and translation products accumulate to very low levels. Consistently with the observations, wsl shows a strong defect in the splicing of chloroplast transcript rpl2, resulting in aberrant transcript accumulation and its product reduction in the mutant. The wsl shows enhanced sensitivity to ABA, salinity, and sugar, and it accumulates more H2O2 than wild-type. These results suggest the reduced translation efficiency may affect the response of the mutant to abiotic stress.
文摘RNA editing sites in plant mitochondria and plastids are addressed by pentatricopeptide repeat （PPR） proteins with E or E and DYW domains, which recognize a specific nucleotide motif upstream of the edited nucleotide. In addition, some sites require MORF proteins for efficient RNA editing. Here, we assign the novel E domain-containing PPR protein, MEF13, as being required for editing at eight sites in Arabidopsis thaliana. A SNP in ecotype C24 altering the editing level at only one of the eight target sites was located by genomic mapping. An EMS mutant allele of the gene for MEF13 was identified in a SNaPshot screen of a mutated plant population. At all eight target sites of MEF13, editing levels are reduced in both morf3 and morf8 mutants, but at only one site in morfl mutants, suggesting that specific MEF13-MORF interactions are required. Yeast two-hybrid analyses detect solid connections of MEF13 with MORF1 and weak contact with MORF3 proteins. Yeast three-hybrid （Y3H） analysis shows that the presence of MORF8 enhances the connection between MEF13 and MORF3, suggesting that a MORF3-MORF8 heteromer may form stably or transiently to establish interaction with MEF13.
基金This work was supported by the Natural Science Foundation of Zhejiang province(LR20C13002)the special support plan for high level talents in Zhejiang Province(2019R52032)Agricultural Sciences and Technologies Innovation Program of the Chinese Academy of Agricultural Sciences.
文摘Pentatricopeptide repeat(PPR)proteins play important roles in the post-transcriptional modification of organellar RNAs in plants.However,the function of most PPR proteins remains unknown.Here,we characterized the rice(Oryza sativa L.)chlorophyll deficient 4(cde4)mutant which exhibits an albino phenotype during early leaf development,with decreased chlorophyll contents and abnormal chloroplasts at low-temperature(20℃).Positional cloning revealed that CDE4 encodes a P-type PPR protein localized in chloroplasts.In the cde4 mutant,plastid-encoded polymerase(PEP)-dependent transcript levels were significantly reduced,but transcript levels of nuclear-encoded genes were increased compared to wild-type plants at 20℃.CDE4 directly binds to the transcripts of the chloroplast genes rpl2,ndhA,and ndhB.Intron splicing of these transcripts was defective in the cde4 mutant at 20℃,but was normal at 32℃.Moreover,CDE4 interacts with the guanylate kinase VIRESCENT 2(V2);overexpression of V2 enhanced CDE4 protein stability,thereby rescuing the cde4 phenotype at 20℃.Our results suggest that CDE4 participates in plastid RNA splicing and plays an important role in rice chloroplast development under lowtemperature conditions.
