Garlic is cultivated worldwide for the value of its bulbs, but its cultivation is challenged by t... more Garlic is cultivated worldwide for the value of its bulbs, but its cultivation is challenged by the infertility of commercial cultivars and the accumulation of pathogens over time, which occurs as a consequence of vegetative (clonal) propagation. In this review, we summarize the state of the art of garlic genetics and genomics, highlighting recent developments that will lead to its development as a modern crop, including the restoration of sexual reproduction in some garlic strains. The set of tools available to the breeder currently includes a chromosome-scale assembly of the garlic genome and multiple transcriptome assemblies that are furthering our understanding of the molecular processes underlying important traits like the infertility, the induction of flowering and bulbing, the organoleptic properties and resistance to various pathogens.
We report the molecular characterization of an ethyl methanesulfonate (EMS)-induced mutation that... more We report the molecular characterization of an ethyl methanesulfonate (EMS)-induced mutation that causes albinism and lethality at the seedling stage in Arabidopsis thaliana. We identified the mutation using a mapping-by-sequencing approach that uses Fisher’s exact tests to detect changes in allele frequencies among the seedlings of an F2 mapping population, which had been pooled according to their phenotypes (wild-type or mutant). After purifying genomic DNA from the plants of both pools, the two samples were sequenced using the Illumina HiSeq 2500 next-generation sequencing platform. The bioinformatic analysis allowed us to identify a point mutation that damages a conserved residue at the acceptor site of an intron of the At2g04030 gene, which encodes the chloroplast-localized AtHsp90.5 protein, a member of the HSP90 family of heat shock proteins. Our RNA-seq analysis demonstrates that the new allele alters the splicing of At2g04030 transcripts in multiple ways, leading to massive...
The International Journal of Developmental Biology, 2001
One of the most frequently represented phenotypic classes among the leaf morphological mutants av... more One of the most frequently represented phenotypic classes among the leaf morphological mutants available in Arabidopsis thaliana is that named Incurvata (Icu), whose members display curled, involute leaves. Here we describe ethyl methanesulfonate-induced alleles of the INCURVATA9 (ICU9) gene, which cause several leaf abnormalities, such as organ radialization and the presence of abaxial trichomes in juvenile leaves. These morphological aberrations can be interpreted as a consequence of a partial transformation of ventral into dorsal leaf identity, which makes the ICU9 gene a candidate for being involved in the specification and/or maintenance of leaf dorsoventrality. In order to identify genetic operations at work during leaf morphogenesis, we have obtained double mutants involving alleles of ten ICU loci, finding that ICU9is functionally related to other five ICU genes. All these six genes seem to be essential for the leaf to properly develop.
The International Journal of Developmental Biology, 2001
In an attempt to identify genes involved in leaf morphogenesis in Arabidopsis thaliana, we screen... more In an attempt to identify genes involved in leaf morphogenesis in Arabidopsis thaliana, we screened for new mutants showing abnormal leaves and conducted genetic analyses of already obtained mutants. Our large-scale mutant search, which got close to but did not reach saturation of the genome, showed that the lines obtained fell into 94 complementation groups. Many of these genes were mapped using a high-throughput linkage analysis method, based on the simultaneous PCR coamplification of 21 polymorphic microsatellites and the fluorescent semiautomated detection of their products. In addition, in an attempt to ascertain wether intraspecific variability might be a source of information on the genetic controls underlying plant leaf morphogenesis, we analyzed variations in the architecture of vegetative leaves in a large sample of Arabidopsis thaliana natural races, concluding that such morphological traits are unlikely to develop under monogenic controls. Hence, a mapping population of ...
Here, we describe how the semidominant, gain-of-function icu4-1 and icu4-2 alleles of the INCURVA... more Here, we describe how the semidominant, gain-of-function icu4-1 and icu4-2 alleles of the INCURVATA4 (ICU4) gene alter leaf phyllotaxis and cell organization in the root apical meristem, reduce root length, and cause xylem overgrowth in the stem. The ICU4 gene was positionally cloned and found to encode the ATHB15 transcription factor, a class III homeodomain/leucine zipper family member, recently named CORONA. The icu4-1 and icu4-2 alleles bear the same point mutation that affects the microRNA complementarity site of ICU4 and is identical to those of several semidominant alleles of the class III homeodomain/leucine zipper family members PHABULOSA and PHAVOLUTA. The icu4-1 and icu4-2 mutations significantly increase leaf transcript levels of the ICU4 gene. The null hst-1 allele of the HASTY gene, which encodes a nucleocytoplasmic transporter, synergistically interacts with icu4-1, the double mutant displaying partial adaxialization of rosette leaves and carpels. Our results suggest ...
Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable conseq... more Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable consequence of the existence of the proximodistal and dorsoventral axes. No gene has been described that affects leaf bilateral symmetry but not dorsoventrality in Arabidopsis thaliana. We screened for viable insertional mutations that affect leaf morphology, and out of more than 700 mutants found only one, desigual1-1 (deal1-1), that exhibited bilateral symmetry breaking but no obvious defects in dorsoventrality. We found that deal1-1 is an allele of VASCULATURE COMPLEXITY AND CONNECTIVITY (VCC). Several overlapping regulatory pathways establish the interspersed lobes and indentations along the margin of Arabidopsis thaliana leaves. These pathways involve feedback loops of auxin, the PIN-FORMED1 (PIN1) auxin efflux carrier, and the CUP-SHAPED COTYLEDON2 (CUC2) transcriptional regulator. Early vcc (deal1) leaf primordia fail to acquire bilateral symmetry and instead form ectopic lobes and sinu...
MicroRNAs (miRNAs) act as negative regulators of gene expression in eukaryotes, a discovery that ... more MicroRNAs (miRNAs) act as negative regulators of gene expression in eukaryotes, a discovery that has opened an expanding field of biological research. Plant miRNAs are known to repress gene expression posttranscriptionally, mainly by guiding cleavage but also by attenuating the translation of target transcripts. In addition, it has been shown that plant miRNAs can also act at the transcriptional level by directing the methylation of target chromosomal loci. Genetic and biochemical approaches are quickly broadening our knowledge of the biogenesis and function of plant miRNAs. Computational approaches have uncovered an unexpectedly large number of miRNAs and their targets in plants. The targets of plant miRNAs often belong to families of transcription factors involved in the control of developmental processes. We review the status of research in this dynamic field, summarizing recent advances in our understanding of the biogenesis and mechanism of action of plant miRNAs, as well as in...
Functional redundancy often hampers the analysis of gene families. To overcome this difficulty, w... more Functional redundancy often hampers the analysis of gene families. To overcome this difficulty, we constructed Arabidopsis thaliana lines that expressed artificial microRNAs designed to simultaneously target two to six paralogous genes encoding members of transcription factor families. Of the 576 genes that we chose as targets, only 122 had already been functionally studied at some level. As a simple indicator of the inhibitory effects of our amiRNAs on their targets, we examined the amiRNA-expressing transgenic lines for morphological phenotypes at the rosette stage. Of 338 transgenes tested, 21 caused a visible morphological phenotype in leaves, a proportion that is much higher than that expected as a result of insertional mutagenesis. Also, our collection probably represents many other mutant phenotypes, not just those in leaves. This robust, versatile method enables functional examination of redundant transcription factor paralogs, and is particularly useful for genes that occur...
We report the complete nucleotide sequence of a new member of the Potyviridae family isolated fro... more We report the complete nucleotide sequence of a new member of the Potyviridae family isolated from passion fruit plants grown in Israel, called Passiflora edulis symptomless virus (PeSV). The PeSV genome is 9,928 nucleotides long and encodes a 3,173 amino acids polyprotein that is predicted to be proteolytically cleaved into 10 mature peptides. Our phylogenetic analysis shows that PeSV represents a new species, and is most closely related to rose yellow mosaic virus (RoYMV). According to currently accepted criteria for genus demarcation, both viruses should be assigned as representative isolates of new species in the recently approved genus, Roymovirus, in the Potyviridae family.
Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable conseq... more Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable consequence of the existence of the proximodistal and dorsoventral axes. No gene has been described that affects leaf bilateral symmetry but not dorsoventrality in Arabidopsis thaliana. We screened for viable insertional mutations that affect leaf morphology, and out of more than 700 mutants found only one, desigual1-1 (deal1-1), that exhibited bilateral symmetry breaking but no obvious defects in dorsoventrality. We found that deal1-1 is an allele of VASCULATURE COMPLEXITY AND CONNECTIVITY (VCC). Several overlapping regulatory pathways establish the interspersed lobes and indentations along the margin of Arabidopsis thaliana leaves. These pathways involve feedback loops of auxin, the PIN-FORMED1 (PIN1) auxin efflux carrier, and the CUP-SHAPED COTYLEDON2 (CUC2) transcriptional regulator. Early vcc (deal1) leaf primordia fail to acquire bilateral symmetry and instead form ectopic lobes and sinu...
MicroRNAs (miRNAs) are ~22-nt single-stranded noncoding RNAs with regulatory roles in a wide rang... more MicroRNAs (miRNAs) are ~22-nt single-stranded noncoding RNAs with regulatory roles in a wide range of cellular functions by repressing eukaryotic gene expression at a post-transcriptional level. Here, we analyzed the effects on meiosis and fertility of hypomorphic or null alleles of the HYL1, HEN1, DCL1, HST and AGO1 genes, which encode miRNA-machinery components in Arabidopsis. Reduced pollen and megaspore mother cell number and fertility were shown by the mutants analyzed. These mutants also exhibited a relaxed chromatin conformation in male meiocytes at the first meiotic division, and increased chiasma frequency, which is likely to be due to increased levels of mRNAs from key genes involved in homologous recombination. The hen1-13 mutant was found to be hypersensitive to gamma irradiation, which mainly causes double-strand breaks susceptible to be repaired by homologous recombination. Our findings uncover a role for miRNA-machinery components in Arabidopsis meiosis, as well as in...
We isolated Arabidopsis thaliana mutants with incurved vegetative leaves. Positional cloning of i... more We isolated Arabidopsis thaliana mutants with incurved vegetative leaves. Positional cloning of incurvata8 (icu8), icu9 and icu15 has identified them as new loss-of-function alleles of the HYPONASTIC LEAVES1 (HYL1), ARGONAUTE1 (AGO1) and HUA ENHANCER1 (HEN1) genes, respectively, which encode known components of the microRNA pathway. The morphological and histological characterization of these mutants and of dicer-like1-9 indicates that small RNAs participate in the proximal-distal and adaxial-abaxial patterning of leaves, as well as in stomatal number establishment. The abnormal vasculature of ago1 and hyl1 leaves also suggests a role for AGO1 and HYL1 in venation patterning. Our mutants expand the allelic series of AGO1, HYL1 and HEN1, and might help to understand the developmental and cellular significance of miRNA-mediated posttranscriptional regulation.
The Plant journal : for cell and molecular biology, 2014
The study and eventual manipulation of leaf development in plants requires a thorough understandi... more The study and eventual manipulation of leaf development in plants requires a thorough understanding of the genetic basis of leaf organogenesis. Forward genetic screens have identified hundreds of Arabidopsis mutants with altered leaf development, but the genome has not yet been saturated. To identify genes required for leaf development we are screening the Arabidopsis Salk Unimutant collection. We have identified 608 lines that exhibit a leaf phenotype with full penetrance and almost constant expressivity and 98 additional lines with segregating mutant phenotypes. To allow indexing and integration with other mutants, the mutant phenotypes were described using a custom leaf phenotype ontology. We found that the indexed mutation is present in the annotated locus for 78% of the 553 mutants genotyped, and that in half of these the annotated T-DNA is responsible for the phenotype. To quickly map non-annotated T-DNA insertions, we developed a reliable, cost-effective and easy method based...
Garlic is cultivated worldwide for the value of its bulbs, but its cultivation is challenged by t... more Garlic is cultivated worldwide for the value of its bulbs, but its cultivation is challenged by the infertility of commercial cultivars and the accumulation of pathogens over time, which occurs as a consequence of vegetative (clonal) propagation. In this review, we summarize the state of the art of garlic genetics and genomics, highlighting recent developments that will lead to its development as a modern crop, including the restoration of sexual reproduction in some garlic strains. The set of tools available to the breeder currently includes a chromosome-scale assembly of the garlic genome and multiple transcriptome assemblies that are furthering our understanding of the molecular processes underlying important traits like the infertility, the induction of flowering and bulbing, the organoleptic properties and resistance to various pathogens.
We report the molecular characterization of an ethyl methanesulfonate (EMS)-induced mutation that... more We report the molecular characterization of an ethyl methanesulfonate (EMS)-induced mutation that causes albinism and lethality at the seedling stage in Arabidopsis thaliana. We identified the mutation using a mapping-by-sequencing approach that uses Fisher’s exact tests to detect changes in allele frequencies among the seedlings of an F2 mapping population, which had been pooled according to their phenotypes (wild-type or mutant). After purifying genomic DNA from the plants of both pools, the two samples were sequenced using the Illumina HiSeq 2500 next-generation sequencing platform. The bioinformatic analysis allowed us to identify a point mutation that damages a conserved residue at the acceptor site of an intron of the At2g04030 gene, which encodes the chloroplast-localized AtHsp90.5 protein, a member of the HSP90 family of heat shock proteins. Our RNA-seq analysis demonstrates that the new allele alters the splicing of At2g04030 transcripts in multiple ways, leading to massive...
The International Journal of Developmental Biology, 2001
One of the most frequently represented phenotypic classes among the leaf morphological mutants av... more One of the most frequently represented phenotypic classes among the leaf morphological mutants available in Arabidopsis thaliana is that named Incurvata (Icu), whose members display curled, involute leaves. Here we describe ethyl methanesulfonate-induced alleles of the INCURVATA9 (ICU9) gene, which cause several leaf abnormalities, such as organ radialization and the presence of abaxial trichomes in juvenile leaves. These morphological aberrations can be interpreted as a consequence of a partial transformation of ventral into dorsal leaf identity, which makes the ICU9 gene a candidate for being involved in the specification and/or maintenance of leaf dorsoventrality. In order to identify genetic operations at work during leaf morphogenesis, we have obtained double mutants involving alleles of ten ICU loci, finding that ICU9is functionally related to other five ICU genes. All these six genes seem to be essential for the leaf to properly develop.
The International Journal of Developmental Biology, 2001
In an attempt to identify genes involved in leaf morphogenesis in Arabidopsis thaliana, we screen... more In an attempt to identify genes involved in leaf morphogenesis in Arabidopsis thaliana, we screened for new mutants showing abnormal leaves and conducted genetic analyses of already obtained mutants. Our large-scale mutant search, which got close to but did not reach saturation of the genome, showed that the lines obtained fell into 94 complementation groups. Many of these genes were mapped using a high-throughput linkage analysis method, based on the simultaneous PCR coamplification of 21 polymorphic microsatellites and the fluorescent semiautomated detection of their products. In addition, in an attempt to ascertain wether intraspecific variability might be a source of information on the genetic controls underlying plant leaf morphogenesis, we analyzed variations in the architecture of vegetative leaves in a large sample of Arabidopsis thaliana natural races, concluding that such morphological traits are unlikely to develop under monogenic controls. Hence, a mapping population of ...
Here, we describe how the semidominant, gain-of-function icu4-1 and icu4-2 alleles of the INCURVA... more Here, we describe how the semidominant, gain-of-function icu4-1 and icu4-2 alleles of the INCURVATA4 (ICU4) gene alter leaf phyllotaxis and cell organization in the root apical meristem, reduce root length, and cause xylem overgrowth in the stem. The ICU4 gene was positionally cloned and found to encode the ATHB15 transcription factor, a class III homeodomain/leucine zipper family member, recently named CORONA. The icu4-1 and icu4-2 alleles bear the same point mutation that affects the microRNA complementarity site of ICU4 and is identical to those of several semidominant alleles of the class III homeodomain/leucine zipper family members PHABULOSA and PHAVOLUTA. The icu4-1 and icu4-2 mutations significantly increase leaf transcript levels of the ICU4 gene. The null hst-1 allele of the HASTY gene, which encodes a nucleocytoplasmic transporter, synergistically interacts with icu4-1, the double mutant displaying partial adaxialization of rosette leaves and carpels. Our results suggest ...
Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable conseq... more Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable consequence of the existence of the proximodistal and dorsoventral axes. No gene has been described that affects leaf bilateral symmetry but not dorsoventrality in Arabidopsis thaliana. We screened for viable insertional mutations that affect leaf morphology, and out of more than 700 mutants found only one, desigual1-1 (deal1-1), that exhibited bilateral symmetry breaking but no obvious defects in dorsoventrality. We found that deal1-1 is an allele of VASCULATURE COMPLEXITY AND CONNECTIVITY (VCC). Several overlapping regulatory pathways establish the interspersed lobes and indentations along the margin of Arabidopsis thaliana leaves. These pathways involve feedback loops of auxin, the PIN-FORMED1 (PIN1) auxin efflux carrier, and the CUP-SHAPED COTYLEDON2 (CUC2) transcriptional regulator. Early vcc (deal1) leaf primordia fail to acquire bilateral symmetry and instead form ectopic lobes and sinu...
MicroRNAs (miRNAs) act as negative regulators of gene expression in eukaryotes, a discovery that ... more MicroRNAs (miRNAs) act as negative regulators of gene expression in eukaryotes, a discovery that has opened an expanding field of biological research. Plant miRNAs are known to repress gene expression posttranscriptionally, mainly by guiding cleavage but also by attenuating the translation of target transcripts. In addition, it has been shown that plant miRNAs can also act at the transcriptional level by directing the methylation of target chromosomal loci. Genetic and biochemical approaches are quickly broadening our knowledge of the biogenesis and function of plant miRNAs. Computational approaches have uncovered an unexpectedly large number of miRNAs and their targets in plants. The targets of plant miRNAs often belong to families of transcription factors involved in the control of developmental processes. We review the status of research in this dynamic field, summarizing recent advances in our understanding of the biogenesis and mechanism of action of plant miRNAs, as well as in...
Functional redundancy often hampers the analysis of gene families. To overcome this difficulty, w... more Functional redundancy often hampers the analysis of gene families. To overcome this difficulty, we constructed Arabidopsis thaliana lines that expressed artificial microRNAs designed to simultaneously target two to six paralogous genes encoding members of transcription factor families. Of the 576 genes that we chose as targets, only 122 had already been functionally studied at some level. As a simple indicator of the inhibitory effects of our amiRNAs on their targets, we examined the amiRNA-expressing transgenic lines for morphological phenotypes at the rosette stage. Of 338 transgenes tested, 21 caused a visible morphological phenotype in leaves, a proportion that is much higher than that expected as a result of insertional mutagenesis. Also, our collection probably represents many other mutant phenotypes, not just those in leaves. This robust, versatile method enables functional examination of redundant transcription factor paralogs, and is particularly useful for genes that occur...
We report the complete nucleotide sequence of a new member of the Potyviridae family isolated fro... more We report the complete nucleotide sequence of a new member of the Potyviridae family isolated from passion fruit plants grown in Israel, called Passiflora edulis symptomless virus (PeSV). The PeSV genome is 9,928 nucleotides long and encodes a 3,173 amino acids polyprotein that is predicted to be proteolytically cleaved into 10 mature peptides. Our phylogenetic analysis shows that PeSV represents a new species, and is most closely related to rose yellow mosaic virus (RoYMV). According to currently accepted criteria for genus demarcation, both viruses should be assigned as representative isolates of new species in the recently approved genus, Roymovirus, in the Potyviridae family.
Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable conseq... more Most plant leaves exhibit bilateral symmetry, which has been hypothesized as an inevitable consequence of the existence of the proximodistal and dorsoventral axes. No gene has been described that affects leaf bilateral symmetry but not dorsoventrality in Arabidopsis thaliana. We screened for viable insertional mutations that affect leaf morphology, and out of more than 700 mutants found only one, desigual1-1 (deal1-1), that exhibited bilateral symmetry breaking but no obvious defects in dorsoventrality. We found that deal1-1 is an allele of VASCULATURE COMPLEXITY AND CONNECTIVITY (VCC). Several overlapping regulatory pathways establish the interspersed lobes and indentations along the margin of Arabidopsis thaliana leaves. These pathways involve feedback loops of auxin, the PIN-FORMED1 (PIN1) auxin efflux carrier, and the CUP-SHAPED COTYLEDON2 (CUC2) transcriptional regulator. Early vcc (deal1) leaf primordia fail to acquire bilateral symmetry and instead form ectopic lobes and sinu...
MicroRNAs (miRNAs) are ~22-nt single-stranded noncoding RNAs with regulatory roles in a wide rang... more MicroRNAs (miRNAs) are ~22-nt single-stranded noncoding RNAs with regulatory roles in a wide range of cellular functions by repressing eukaryotic gene expression at a post-transcriptional level. Here, we analyzed the effects on meiosis and fertility of hypomorphic or null alleles of the HYL1, HEN1, DCL1, HST and AGO1 genes, which encode miRNA-machinery components in Arabidopsis. Reduced pollen and megaspore mother cell number and fertility were shown by the mutants analyzed. These mutants also exhibited a relaxed chromatin conformation in male meiocytes at the first meiotic division, and increased chiasma frequency, which is likely to be due to increased levels of mRNAs from key genes involved in homologous recombination. The hen1-13 mutant was found to be hypersensitive to gamma irradiation, which mainly causes double-strand breaks susceptible to be repaired by homologous recombination. Our findings uncover a role for miRNA-machinery components in Arabidopsis meiosis, as well as in...
We isolated Arabidopsis thaliana mutants with incurved vegetative leaves. Positional cloning of i... more We isolated Arabidopsis thaliana mutants with incurved vegetative leaves. Positional cloning of incurvata8 (icu8), icu9 and icu15 has identified them as new loss-of-function alleles of the HYPONASTIC LEAVES1 (HYL1), ARGONAUTE1 (AGO1) and HUA ENHANCER1 (HEN1) genes, respectively, which encode known components of the microRNA pathway. The morphological and histological characterization of these mutants and of dicer-like1-9 indicates that small RNAs participate in the proximal-distal and adaxial-abaxial patterning of leaves, as well as in stomatal number establishment. The abnormal vasculature of ago1 and hyl1 leaves also suggests a role for AGO1 and HYL1 in venation patterning. Our mutants expand the allelic series of AGO1, HYL1 and HEN1, and might help to understand the developmental and cellular significance of miRNA-mediated posttranscriptional regulation.
The Plant journal : for cell and molecular biology, 2014
The study and eventual manipulation of leaf development in plants requires a thorough understandi... more The study and eventual manipulation of leaf development in plants requires a thorough understanding of the genetic basis of leaf organogenesis. Forward genetic screens have identified hundreds of Arabidopsis mutants with altered leaf development, but the genome has not yet been saturated. To identify genes required for leaf development we are screening the Arabidopsis Salk Unimutant collection. We have identified 608 lines that exhibit a leaf phenotype with full penetrance and almost constant expressivity and 98 additional lines with segregating mutant phenotypes. To allow indexing and integration with other mutants, the mutant phenotypes were described using a custom leaf phenotype ontology. We found that the indexed mutation is present in the annotated locus for 78% of the 553 mutants genotyped, and that in half of these the annotated T-DNA is responsible for the phenotype. To quickly map non-annotated T-DNA insertions, we developed a reliable, cost-effective and easy method based...
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