Project description:Plant root architecture is a major determinant of fitness, and is under constant modification in response to favorable and unfavorable environmental stimuli. Beyond impacts on the primary root, the environment can also alter the position, spacing, density, and length of secondary or lateral roots. Lateral root development is among the best-studied developmental processes in Arabidopsis thaliana, yet the earliest steps of organogenesis remain elusive. Among the challenges faced in capturing these early molecular events is the fact that this process occurs in a small number of cells with unpredictable timing. The advent of single-cell sequencing affords the opportunity to isolate cells undergoing this fate transition and examine their transcriptomes independently. Using this approach, we successfully captured the transcriptomes of lateral root primordia and discovered many previously unreported upregulated genes. To further study this process, we developed a method to selectively repress genes in the xylem pole pericycle cells where lateral roots originate. We found that expression of several of the upregulated genes was required for normal root development. In addition, we discovered a subpopulation of cells in the endodermal cell file that respond to lateral root initiation, further highlighting the benefits of the single cell approach.
Project description:We performed an analysis of transcriptomic responses to auxin within four distinct tissues of the Arabidopsis thaliana root. This high-resolution dataset shows how different cell types are predisposed to react to auxin with discrete transcriptional responses. The sensitivity provided by the analysis lies in the ability to detect cell-type specific responses diluted in organ-level analyses. This dataset provides a novel resource to examine how auxin, a widespread signal in plant development, influences differentiation and patterning in the plant through tissue-specific transcriptional regulation. To analyze the effect of auxin in separate spatial domains of the root, early transcriptional changes in response to auxin treatment were assayed by means of fluorescence activated cell sorting (FACS) and microarray analysis in four tissues of the Arabidopsis root (wild type Col-0). The samples covered inner and outer as well as proximal and distal cell populations; including the stele (reporter line pWOL::GFP), xylem-pole (xp) pericycle (enhancer trap line E3754), epidermis/lateral root cap (reporter line pWER::GFP) and columella (enhancer trap line PET111). One-week-old seedlings of the individual lines were treated with auxin (two hours, 5µM indole-3-acetic acid [IAA]) or mock treated, after which roots were harvested and cells were dissociated by cell wall digestion (1 hour; including 5uM IAA) . GFP-positive cells were sorted and used for microarray transcriptome analysis (as in Bargmann and Birnbaum, Plant Phys. 2010). For comparison, transcriptional responses to auxin were also assayed in intact (undigested) roots.
Project description:To optimize access to nitrogen under limiting conditions, root systems must continuously sense and respond to local or temporal fluctuations in nitrogen availability. In Arabidopsis thaliana and several other species, external N levels that induce only mild deficiency stimulate the emergence of lateral roots and especially the elongation of primary and lateral roots. However, the identity of the genes involved in this coordination remains still largely elusive. In order to identify novel genes and mechanisms underlying nitrogen-dependent root morphological changes, we investigated time-dependent changes in the root transcriptome of Arabidopsis thaliana plants grown under sufficient nitrogen or under conditions that induced mild nitrogen deficiency.
Project description:Coordinated distribution of Pi between roots and shoots is an important process that plants use to maintain Pi homeostasis. SHR (SHORT-ROOT) is well-characterized for its function in root radial patterning1-3. Here, we demonstrate a new role of SHR in controlling phosphate (Pi) allocation from roots to shoots by regulating PHOSPHATE1 (PHO1) in the root differentiation zone. We recovered a weak mutant allele of SHR in Arabidopsis which accumulates much less Pi in the shoot and shows constitutive Pi starvation response (PSR) under Pi-sufficient condition. Besides, Pi starvation suppresses SHR protein accumulation and releases its inhibition on the HD-ZIP Ⅲ transcription factor PHB. PHB accumulates and directly binds the promoter of PHO2 to upregulate its transcription, resulting in PHO1 degradation in the xylem-pole pericycle cells. Our findings reveal a previously unrecognized mechanism of how plants repress Pi translocation from roots to shoots in response to Pi starvation.
Project description:Phosphate (Pi) deficiency alters root hair length and frequency as a means of increasing the absorptive surface area of roots. Three partly redundant single R3 MYB proteins, CAPRICE (CPC), ENHANCER OF TRY AND CPC1 (ETC1) and TRIPTYCHON (TRY), positively regulate the root hair cell fate by participating in a lateral inhibition mechanism. To identify putative targets and processes that are controlled by these three transcription factors (TFs), we conducted transcriptional profiling of roots from Arabidopsis thaliana wild-type plants, and cpc, etc1 and try mutants grown under Pi-replete and Pi-deficient conditions using RNA-seq.
Project description:Arabidopsis thaliana plants were grown from seeds in Petri dishes on MS medium. 4 days old plants were co-incubated with L. bicolor without physical contact. After 2 days of co-incubation, roots were either harvested whole or separated into the root tip (ca. 1/4 of the whole root) and the lateral root zone (= remaining root).
Project description:De novo shoot organogenesis (DNSO) is a commonly used pathway for plant biotechnology, and is a hormonally regulated process, where auxin and cytokinin coordinates suites of genes encoding transcription factors, general transcription factors, and RNA metabolism machinery genes. Here we report that silencing Arabidopsis thaliana CTD phosphatase-like 4 (CPL4RNAi), which increases phosphorylation level of RNA polymerase II (pol II) CTD, altered lateral root development and DNSO efficiency of the host plants, suggesting an importance of precise control of pol II activities during DNSO. Under standard condition, roots of CPL4RNAi lines produced no or few lateral roots. When induced by high concentration of auxin, CPL4RNAi lines failed to produce focused auxin maxima at the meristem of lateral root primordia, and produced fasciated lateral roots. By contrast, root explants of CPL4RNAi lines were highly competent for DNSO. Efficient DNSO of CPL4RNAi lines were observed even under 10 times less cytokinin required for wild type explants. Transcriptome analysis showed CPL4RNAi but not wild type explants expressed high levels of shoot meristem related genes during priming by high auxin/cytokinin ratio, and subsequent shoot induction with cytokinin. These results indicate that CPL4 functions as a repressor of the early stage of DNSO, during acquisition of competency by high auxin/cytokinin ratio, perhaps via regulation of pol II activities.
Project description:Transcriptional profiling of Arabidopsis thalina seedlings in response to high (140 µM) and low (20 µM) concentrations of chromate (K2CrO4). Low concentrations of chromate (e.g. 40 µM) promoted primary root growth, while high concentrations (e.g. 140 µM) repressed growth and increased formation of root hairs, lateral roots and adventitious roots.
Project description:When grown under phosphate (Pi) deficiency, plants adjust their developmental program and metabolic activity to cope with this nutritional stress. For Arabidopsis, the developmental responses include inhibition of primary root growth and enhanced formation of lateral roots and root hairs. Pi deficiency also inhibits photosynthesis by suppressing the expression of photosynthetic genes. Interestingly, early studies showed that photosynthetic gene expression was also suppressed in roots, a non-photosynthetic tissue. The biological relevance of this phenomenon, however, is not known. In this work, we characterized an Arabidopsis mutant, hps7, which is hypersensitive to Pi deficiency; the hypersensitivity includes an increased inhibition of root growth. HPS7 encodes a tyrosylprotein sulfotransferase (TPST). Accumulation of TPST proteins, but not mRNA, is induced by Pi deficiency. Comparative RNA-Seq analyses indicated that expression of many photosynthetic genes was activated in the roots of hps7. Under Pi deficiency, the expression of the photosynthetic genes in hps7 is further increased, which leads to the enhanced accumulation of chlorophyll, starch, and reactive oxygen species. The increased inhibition of root growth in hps7 under Pi deficiency was completely reversed by growing plants in the dark. Based on these results, we propose that suppression of photosynthetic gene expression in roots is required for sustained root growth under Pi deficiency.