ORCID Profile
0000-0003-3299-5287
Current Organisation
University of Oxford
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Publisher: Oxford University Press (OUP)
Date: 25-04-2019
DOI: 10.1105/TPC.19.00132
Publisher: Cold Spring Harbor Laboratory
Date: 02-02-2021
DOI: 10.1101/2021.02.02.429330
Abstract: The Arabidopsis gene Chloroplast Import Apparatus 2 (CIA2) encodes a transcription factor that positively affects the activity of nuclear genes for chloroplast ribosomal proteins and chloroplast protein import machineries. CIA2-like (CIL) is the paralogous gene of CIA2 . We generated a cil mutant by site-directed mutagenesis and compared it with cia2 and cia2cil double mutant. Phenotype of the cil mutant did not differ from the wild type under our growth conditions, except faster growth and earlier time to flowering. Compared to cia2, the cia2cil mutant showed more impaired chloroplast functions and reduced amounts of plastid ribosomal RNAs. In silico analyses predict for CIA2 and CIL a C-terminal CCT domain and an N-terminal chloroplast transit peptide (cTP). Chloroplast (and potentially nuclear) localization was previously shown for HvCMF3 and HvCMF7, the homologs of CIA2 and CIL in barley. We observed nuclear localization of CIL after transient expression in Arabidopsis protoplasts. Surprisingly, transformation of cia2 with HvCMF3, HvCMF7 or with a truncated CIA2 lacking the predicted cTP could partially rescue the pale-green phenotype of cia2 . These data are discussed with respect to potentially overlapping functions between CIA2, CIL and their barley homologs and to the function of the putative cTPs of CIA2 and CIL. The nucleus-localized CCT domain proteins CIA2 and CIL in Arabidopsis and the homologous chloroplast-localized HvCMF3 and HvCMF7 in barley retained partially overlapping functions in chloroplast development.
Publisher: Cold Spring Harbor Laboratory
Date: 04-02-2021
DOI: 10.1101/2021.02.04.429718
Abstract: Implementation of next-generation sequencing in forward genetic screens greatly accelerated gene discovery in species with larger genomes, including many crop plants. In barley, extensive mutant collections are available, however, the causative mutations for many of the genes remains largely unknown. Here we demonstrate how a combination of low-resolution genetic mapping, whole-genome resequencing and comparative functional analyses provides a promising path towards candidate identification of genes involved in plastid biology and / or photosynthesis, even if genes are located in recombination poor regions of the genome. As a proof of concept, we simulated the prediction of a candidate gene for the recently cloned variegation mutant albostrians ( HvAST / HvCMF7 ) and adopted the approach for suggesting HvClpC1 as candidate gene for the yellow-green variegation mutant luteostrians . Forward genetics is an approach of identifying a causal gene for a mutant phenotype and has proven to be a powerful tool for dissecting the genetic control of biological processes in many species. A large number of barley mutants was generated in the 1940s to 1970s when mutation breeding programs flourished. Genetic dissection of the causative mutations responsible for the phenotype, however, lagged far behind, limited by lack of molecular markers and high-throughput genotyping platforms. Next-generation sequencing technologies have revolutionized genomics, facilitating the process of identifying mutations underlying a phenotype of interest. Multiple mapping-by-sequencing or cloning-by-sequencing strategies were established towards fast gene discovery. In this study, we used mapping-by-sequencing to identify candidate genes within coarsely delimited genetic intervals, for two variegation mutants in barley – luteostrians and albostrians . After testing the approach using the ex le of the previously cloned albostrians gene HvAST , the gene HvClpC1 could be delimited as candidate gene for luteostrians. The mapping-by-sequencing strategy implemented here is generally suited for surveying barley mutant collections for phenotypes affecting fundamental processes of plant morphology, physiology and development.
Publisher: Frontiers Media SA
Date: 07-06-2021
Abstract: The Arabidopsis gene Chloroplast Import Apparatus 2 ( CIA2 ) encodes a transcription factor that positively affects the activity of nuclear genes for chloroplast ribosomal proteins and chloroplast protein import machineries. CIA2-like ( CIL ) is the paralogous gene of CIA2 . We generated a cil mutant by site-directed mutagenesis and compared it with cia2 and cia2cil double mutant. Phenotype of the cil mutant did not differ from the wild type under our growth conditions, except faster growth and earlier time to flowering. Compared to cia2 , the cia2cil mutant showed more impaired chloroplast functions and reduced amounts of plastid ribosomal RNAs. In silico analyses predict for CIA2 and CIL a C-terminal CCT domain and an N-terminal chloroplast transit peptide (cTP). Chloroplast (and potentially nuclear) localization was previously shown for HvCMF3 and HvCMF7, the homologs of CIA2 and CIL in barley. We observed nuclear localization of CIL after transient expression in Arabidopsis protoplasts. Surprisingly, transformation of cia2 with HvCMF3 , HvCMF7 , or with a truncated CIA2 lacking the predicted cTP could partially rescue the pale-green phenotype of cia2 . These data are discussed with respect to potentially overlapping functions between CIA2, CIL, and their barley homologs and to the function of the putative cTPs of CIA2 and CIL.
Publisher: Cold Spring Harbor Laboratory
Date: 09-09-2019
DOI: 10.1101/756833
Abstract: Gene pairs resulting from whole genome duplication (WGD), so-called ohnologous genes, are retained only if at least one gene of the pair undergoes neo- or subfunctionalization. Sequence-based phylogenetic analyses of the ohnologous genes ALBOSTRIANS ( HvAST/HvCMF7 ) and A LBO S TRIANS- L IKE ( HvASL / HvCMF3 ) of barley ( Hordeum vulgare ) revealed that they belong to a newly identified subfamily of genes encoding CCT domain proteins with putative N-terminal chloroplast transit peptides. Recently, we showed that HvCMF7 is needed for chloroplast ribosome biogenesis. Here we demonstrate that mutations in HvCMF3 lead to seedlings delayed in development. They exhibit a xantha phenotype and successively develop pale green leaves. Compared to the wild type, plastids of the mutant seedlings show decreased PSII efficiency and lower amounts of ribosomal RNAs they contain less thylakoids and grana with a higher number of more loosely stacked thylakoid membranes. Site-directed mutagenesis of HvCMF3 identified a previously unknown functional region, which is highly conserved within this subfamily of CCT domain containing proteins. HvCMF3:GFP fusion constructs localized to plastids. Hvcmf3Hvcmf7 double mutants indicated epistatic activity of HvCMF7 over HvCMF3. The chloroplast ribosome deficiency is discussed as the primary defect of the Hvcmf3 mutants. Our data suggests that HvCMF3 and HvCMF7 have similar but not identical functions. Phylogenetic and mutant analyses of the barley protein HvCMF3 (ALBOSTRIANS-LIKE) identified, in higher plants, a subfamily of CCT domain proteins with essential function in chloroplast development.
Publisher: Frontiers Media SA
Date: 10-2021
Abstract: Gene pairs resulting from whole genome duplication (WGD), so-called ohnologous genes, are retained if at least one member of the pair undergoes neo- or sub-functionalization. Phylogenetic analyses of the ohnologous genes ALBOSTRIANS ( HvAST/HvCMF7 ) and A LBO S TRIANS- L IKE ( HvASL / HvCMF3 ) of barley ( Hordeum vulgare ) revealed them as members of a subfamily of genes coding for CCT motif ( C ONSTANS, C ONSTANS-LIKE and T IMING OF CAB1) proteins characterized by a single CCT domain and a putative N-terminal chloroplast transit peptide. Recently, we showed that HvCMF7 is needed for chloroplast ribosome biogenesis. Here we demonstrate that mutations in HvCMF3 lead to seedlings delayed in development. They exhibit a yellowish/light green – xantha – phenotype and successively develop pale green leaves. Compared to wild type, plastids of mutant seedlings show a decreased PSII efficiency, impaired processing and reduced amounts of ribosomal RNAs they contain less thylakoids and grana with a higher number of more loosely stacked thylakoid membranes. Site-directed mutagenesis of HvCMF3 identified a previously unknown functional domain, which is highly conserved within this subfamily of CCT domain containing proteins. HvCMF3:GFP fusion constructs were localized to plastids and nucleus. Hvcmf3Hvcmf7 double mutants exhibited a xantha -albino or albino phenotype depending on the strength of molecular lesion of the HvCMF7 allele. The chloroplast ribosome deficiency is discussed as the primary observed defect of the Hvcmf3 mutants. Based on our observations, the genes HvCMF3 and HvCMF7 have similar but not identical functions in chloroplast development of barley supporting our hypothesis of neo-/sub-functionalization between both ohnologous genes.
Publisher: Frontiers Media SA
Date: 16-04-2021
Abstract: Implementation of next-generation sequencing in forward genetic screens greatly accelerated gene discovery in species with larger genomes, including many crop plants. In barley, extensive mutant collections are available, however, the causative mutations for many of the genes remains largely unknown. Here we demonstrate how a combination of low-resolution genetic mapping, whole-genome resequencing and comparative functional analyses provides a promising path toward candidate identification of genes involved in plastid biology and/or photosynthesis, even if genes are located in recombination poor regions of the genome. As a proof of concept, we simulated the prediction of a candidate gene for the recently cloned variegation mutant albostrians ( HvAST/HvCMF7 ) and adopted the approach for suggesting HvClpC1 as candidate gene for the yellow-green variegation mutant luteostrians .
Publisher: Cold Spring Harbor Laboratory
Date: 25-02-2019
DOI: 10.1101/560797
Abstract: Chloroplasts fuel plant development and growth by converting solar into chemical energy. They mature from proplastids through the concerted action of genes in both the organellar and the nuclear genome. Defects in such genes impair chloroplast development and may lead to pigment-deficient seedlings or seedlings with variegated leaves. Such mutants are instrumental as tools for dissecting genetic factors underlying the mechanisms involved in chloroplast biogenesis. Characterization of the green-white variegated albostrians mutant of barley has greatly broadened the field of chloroplast biology including the discovery of retrograde signaling. Here, we report the identification of the ALBOSTRIANS gene HvAST by positional cloning as well as its functional validation based on independently induced mutants by TILLING and RNA-guided Cas9 endonuclease mediated gene editing. The phenotypes of the independent HvAST mutants imply residual activity of HvAST in the original albostrians allele conferring an imperfect penetrance of the variegated phenotype even at homozygous state of the mutation. HvAST is a homolog of the Arabidopsis thaliana CCT Motif transcription factor gene AtCIA2 , which was reported to be involved in the expression of nuclear genes essential for chloroplast biogenesis. Interestingly, in barley we localized HvAST to the chloroplast indicating novel without any clear evidence of nuclear localization. Leaf variegation in the barley mutant albostrians is caused by mutation of a single CCT-domain containing gene with residual activity, which is directed to the chloroplast.
Location: United Kingdom of Great Britain and Northern Ireland
Location: Germany
No related grants have been discovered for Mingjiu Li.