Aashish Ranjan

A Quantitative Genetic Basis for Leaf Morphology in a Set of Precisely Defined Tomato Introgression Lines

Natural variation leading to differences in leaf morphology between domesticated tomato and a wild relative is explored in a set of introgression lines. The phenotypic context of leaf morphology with other traits is examined at the whole-plant level, with implications for organ-specific breeding efforts. Introgression lines (ILs), in which genetic material from wild tomato species is introgressed into a domesticated background, have been used extensively in tomato (Solanum lycopersicum) improvement. Here, we genotype an IL population derived from the wild desert tomato Solanum pennellii at ultrahigh density, providing the exact gene content harbored by each line. To take advantage of this information, we determine IL phenotypes for a suite of vegetative traits, ranging from leaf complexity, shape, and size to cellular traits, such as stomatal density and epidermal cell phenotypes. Elliptical Fourier descriptors on leaflet outlines provide a global analysis of highly heritable, intricate aspects of leaf morphology. We also demonstrate constraints between leaflet size and leaf complexity, pavement cell size, and stomatal density and show independent segregation of traits previously assumed to be genetically coregulated. Meta-analysis of previously measured traits in the ILs shows an unexpected relationship between leaf morphology and fruit sugar levels, which RNA-Seq data suggest may be attributable to genetically coregulated changes in fruit morphology or the impact of leaf shape on photosynthesis. Together, our results both improve upon the utility of an important genetic resource and attest to a complex, genetic basis for differences in leaf morphology between natural populations.

De novo assembly and characterization of the transcriptome of the parasitic weed Cuscuta pentagona identifies genes associated with plant parasitism

Transcriptional dynamics during parasitism in the parasitic weed Cuscuta pentagona reveals increased expression of genes encoding transporters and stimulus response regulators, and a decrease in the expression of genes encoding photosynthetic proteins. Parasitic flowering plants are one of the most destructive agricultural pests and have major impact on crop yields throughout the world. Being dependent on finding a host plant for growth, parasitic plants penetrate their host using specialized organs called haustoria. Haustoria establish vascular connections with the host, which enable the parasite to steal nutrients and water. The underlying molecular and developmental basis of parasitism by plants is largely unknown. In order to investigate the process of parasitism, RNAs from different stages (i.e. seed, seedling, vegetative strand, prehaustoria, haustoria, and flower) were used to de novo assemble and annotate the transcriptome of the obligate plant stem parasite dodder (Cuscuta pentagona). The assembled transcriptome was used to dissect transcriptional dynamics during dodder development and parasitism and identified key gene categories involved in the process of plant parasitism. Host plant infection is accompanied by increased expression of parasite genes underlying transport and transporter categories, response to stress and stimuli, as well as genes encoding enzymes involved in cell wall modifications. By contrast, expression of photosynthetic genes is decreased in the dodder infective stages compared with normal stem. In addition, genes relating to biosynthesis, transport, and response of phytohormones, such as auxin, gibberellins, and strigolactone, were differentially expressed in the dodder infective stages compared with stems and seedlings. This analysis sheds light on the transcriptional changes that accompany plant parasitism and will aid in identifying potential gene targets for use in controlling the infestation of crops by parasitic weeds.

De Novo Assembly and Characterization of the Transcriptome of the Parasitic Weed Dodder Identifies Genes Associated with Plant Parasitism

Transcriptional dynamics during parasitism in the parasitic weed Cuscuta pentagona reveals increased expression of genes encoding transporters and stimulus response regulators, and a decrease in the expression of genes encoding photosynthetic proteins. Parasitic flowering plants are one of the most destructive agricultural pests and have major impact on crop yields throughout the world. Being dependent on finding a host plant for growth, parasitic plants penetrate their host using specialized organs called haustoria. Haustoria establish vascular connections with the host, which enable the parasite to steal nutrients and water. The underlying molecular and developmental basis of parasitism by plants is largely unknown. In order to investigate the process of parasitism, RNAs from different stages (i.e. seed, seedling, vegetative strand, prehaustoria, haustoria, and flower) were used to de novo assemble and annotate the transcriptome of the obligate plant stem parasite dodder (Cuscuta pentagona). The assembled transcriptome was used to dissect transcriptional dynamics during dodder development and parasitism and identified key gene categories involved in the process of plant parasitism. Host plant infection is accompanied by increased expression of parasite genes underlying transport and transporter categories, response to stress and stimuli, as well as genes encoding enzymes involved in cell wall modifications. By contrast, expression of photosynthetic genes is decreased in the dodder infective stages compared with normal stem. In addition, genes relating to biosynthesis, transport, and response of phytohormones, such as auxin, gibberellins, and strigolactone, were differentially expressed in the dodder infective stages compared with stems and seedlings. This analysis sheds light on the transcriptional changes that accompany plant parasitism and will aid in identifying potential gene targets for use in controlling the infestation of crops by parasitic weeds.

A Modern Ampelography: A Genetic Basis for Leaf Shape and Venation Patterning in Grape

Statistical methods can globally describe the complex shapes of grape leaves, permitting the evaluation of not only the genetic basis of leaf shape but its correlation with traits of economic interest. Terroir, the unique interaction between genotype, environment, and culture, is highly refined in domesticated grape (Vitis vinifera). Toward cultivating terroir, the science of ampelography tried to distinguish thousands of grape cultivars without the aid of genetics. This led to sophisticated phenotypic analyses of natural variation in grape leaves, which within a palmate-lobed framework exhibit diverse patterns of blade outgrowth, hirsuteness, and venation patterning. Here, we provide a morphometric analysis of more than 1,200 grape accessions. Elliptical Fourier descriptors provide a global analysis of leaf outlines and lobe positioning, while a Procrustes analysis quantitatively describes venation patterning. Correlation with previous ampelography suggests an important genetic component, which we confirm with estimates of heritability. We further use RNA-Seq of mutant varieties and perform a genome-wide association study to explore the genetic basis of leaf shape. Meta-analysis reveals a relationship between leaf morphology and hirsuteness, traits known to correlate with climate in the fossil record and extant species. Together, our data demonstrate a genetic basis for the intricate diversity present in grape leaves. We discuss the possibility of using grape leaves as a breeding target to preserve terroir in the face of anticipated climate change, a major problem facing viticulture.

Domestication selected for deceleration of the circadian clock in cultivated tomato

The circadian clock is a critical regulator of plant physiology and development, controlling key agricultural traits in crop plants. In addition, natural variation in circadian rhythms is important for local adaptation. However, quantitative modulation of circadian rhythms due to artificial selection has not yet been reported. Here we show that the circadian clock of cultivated tomato (Solanum lycopersicum) has slowed during domestication. Allelic variation of the tomato homolog of the Arabidopsis gene EID1 is responsible for a phase delay. Notably, the genomic region harboring EID1 shows signatures of a selective sweep. We find that the EID1 allele in cultivated tomatoes enhances plant performance specifically under long day photoperiods, suggesting that humans selected slower circadian rhythms to adapt the cultivated species to the long summer days it encountered as it was moved away from the equator.

Resolving Distinct Genetic Regulators of Tomato Leaf Shape within a Heteroblastic and Ontogenetic Context

Leaf shape is dynamic and influenced by numerous factors. Here, the authors quantify shape differences in leaves across the leaf series and during their development, determining their genetic basis and the effects of environment, in a population resulting from a cross between tomato and a wild desert relative. Leaf shape is mutable, changing in ways modulated by both development and environment within genotypes. A complete model of leaf phenotype would incorporate the changes in leaf shape during juvenile-to-adult phase transitions and the ontogeny of each leaf. Here, we provide a morphometric description of >33,000 leaflets from a set of tomato (Solanum spp) introgression lines grown under controlled environment conditions. We first compare the shape of these leaves, arising during vegetative development, with >11,000 previously published leaflets from a field setting and >11,000 leaflets from wild tomato relatives. We then quantify the changes in shape, across ontogeny, for successive leaves in the heteroblastic series. Using principal component analysis, we then separate genetic effects modulating (1) the overall shape of all leaves versus (2) the shape of specific leaves in the series, finding the former more heritable than the latter and comparing quantitative trait loci regulating each. Our results demonstrate that phenotype is highly contextual and that unbiased assessments of phenotype, for quantitative genetic or other purposes, would ideally sample the many developmental and environmental factors that modulate it.

The Arabidopsis repressor of light signaling SPA1 acts in the phloem to regulate seedling de-etiolation, leaf expansion and flowering time

Plants adjust their growth and development in response to the ambient light environment. These light responses involve systemic signals that coordinate differentiation of different tissues and organs. Here, we have investigated the function of the key repressor of photomorphogenesis SPA1 in different tissues of the plant by expressing GUS-SPA1 under the control of tissue-specific promoters in a spa mutant background. We show that SPA1 expression in the phloem vasculature is sufficient to rescue the spa1 mutant phenotype in dark-grown spa mutant seedlings. Expression of SPA1 in mesophyll, epidermis or root tissues of the seedling, by contrast, has no or only slight effects. In the leaf, SPA1 expression in both the phloem and the mesophyll is required for full complementation of the defect in leaf expansion. SPA1 in phloem and mesophyll tissues affected division and expansion of cells in the epidermal layer, indicating that SPA1 induces non-cell-autonomous responses also in the leaf. Photoperiodic flowering is exclusively controlled by SPA1 expression in the phloem, which is consistent with previous results showing that the direct substrate of the COP1/SPA complex, CONSTANS, also acts in the phloem. Taken together, our results highlight the importance of phloem vascular tissue in coordinating growth and development. Because the SPA1 protein itself is incapable of moving from cell to cell, we suggest that SPA1 regulates the activity of downstream component(s) of light signaling that subsequently act in a non-cell-autonomous manner. SPA1 action in the phloem may also result in mechanical stimuli that affect cell elongation and cell division in other tissues.

The tomato genome: implications for plant breeding, genomics and evolution

The genome sequence of tomato (Solanum lycopersicum), one of the most important vegetable crops, has recently been decoded. We address implications of the tomato genome for plant breeding, genomics and evolutionary studies, and its potential to fuel future crop biology research.

Auxin represses stomatal development in dark-grown seedlings via Aux/IAA proteins

Stomatal development is tightly regulated through internal and external factors that are integrated by a complex signalling network. Light represents an external factor that strongly promotes stomata formation. Here, we show that auxin-resistant aux/iaa mutants, e.g. axr3-1, exhibit a de-repression of stomata differentiation in dark-grown seedlings. The higher stomatal index in dark-grown axr3-1 mutants when compared with the wild type is due to increased cell division in the stomatal lineage. Excessive stomata in dark-grown seedlings were also observed in mutants defective in auxin biosynthesis or auxin perception and in seedlings treated with the polar auxin transport inhibitor NPA. Consistent with these findings, exogenous auxin repressed stomata formation in light-grown seedlings. Taken together, these results indicate that auxin is a negative regulator of stomatal development in dark-grown seedlings. Epistasis analysis revealed that axr3-1 acts genetically upstream of the bHLH transcription factors SPCH, MUTE and FAMA, as well as the YDA MAP kinase cascade, but in parallel with the repressor of photomorphogenesis COP1 and the receptor-like protein TMM. The effect of exogenous auxin required the ER family of leucine-rich repeat receptor-like kinases, suggesting that auxin acts at least in part through the ER family. Expression of axr3-1 in the stomatal lineage was insufficient to alter the stomatal index, implying that cell-cell communication is necessary to mediate the effect of auxin. In summary, our results show that auxin signalling contributes to the suppression of stomatal differentiation observed in dark-grown seedlings.

Functional analysis of COP1 and SPA orthologs from Physcomitrella and rice during photomorphogenesis of transgenic Arabidopsis reveals distinct evolutionary conservation

BackgroundPlants have evolved light sensing mechanisms to optimally adapt their growth and development to the ambient light environment. The COP1/SPA complex is a key negative regulator of light signaling in the well-studied dicot Arabidopsis thaliana. COP1 and members of the four SPA proteins are part of an E3 ubiquitin ligase that acts in darkness to ubiquitinate several transcription factors involved in light responses, thereby targeting them for degradation by the proteasome. While COP1 is also found in humans, SPA proteins appear specific to plants. Here, we have functionally addressed evolutionary conservation of COP1 and SPA orthologs from the moss Physcomitrella, the monocot rice and the dicot Arabidopsis.ResultsTo this end, we analyzed the activities of COP1- and SPA-like proteins from Physcomitrella patens and rice when expressed in Arabidopsis. Expression of rice COP1 and Physcomitrella COP1 protein sequences predominantly complemented all phenotypic aspects of the viable, hypomorphic cop1-4 mutant and the null, seedling-lethal cop1-5 mutant of Arabidopsis: rice COP1 fully rescued the constitutive-photomorphogenesis phenotype in darkness and the leaf expansion defect of cop1 mutants, while it partially restored normal photoperiodic flowering in cop1. Physcomitrella COP1 partially restored normal seedling growth and flowering time, while it fully restored normal leaf expansion in the cop1 mutants. In contrast, expression of a SPA ortholog from Physcomitrella (PpSPAb) in Arabidopsis spa mutants did not rescue any facet of the spa mutant phenotype, suggesting that the PpSPAb protein is not functionally conserved or that the Arabidopsis function evolved after the split of mosses and seed plants. The SPA1 ortholog from rice (OsSPA1) rescued the spa mutant phenotype in dark-grown seedlings, but did not complement any spa mutant phenotype in light-grown seedlings or in adult plants.ConclusionOur results show that COP1 protein sequences from Physcomitrella, rice and Arabidopsis have been functionally conserved during evolution, while the SPA proteins showed considerable functional divergence. This may - at least in part - reflect the fact that COP1 is a single copy gene in seed plants, while SPA proteins are encoded by a small gene family of two to four members with possibly sub- or neofunctionalized tasks.