Project description:Arabidopsis thaliana plant expressing 35S:WIND1 shows callus-like morphology without hormone treatment. Transcriptomes of the callus-like cell expressing 35S:WIND1, callus of T87 cultured cell, 2,4-D-induced callus and control seedling plant were compared by Agilent microarray.
Project description:Arabidopsis thaliana plant expressing 35S:WIND1 shows callus-like morphology without hormone treatment. Transcriptomes of the callus-like cell expressing 35S:WIND1, callus of T87 cultured cell, 2,4-D-induced callus and control seedling plant were compared by Agilent microarray. Comparison of four kinds of Arabidopsis thaliana plants. Biological replicates: three for each.
Project description:In plant tissue culture, callus forms from detached explants in response to a high-auxin-to-low-cytokinin ratio on callus-inducing medium. Callus is a group of pluripotent cells because it can regenerate either roots or shoots in response to a low level of auxin on root-inducing medium or a high-cytokinin-to-low-auxin ratio on shoot-inducing medium, respectively1. However, our knowledge of the mechanism of pluripotency acquisition during callus formation is limited. On the basis of analyses at the single-cell level, we show that the tissue structure of Arabidopsis thaliana callus on callus-inducing medium is similar to that of the root primordium or root apical meristem, and the middle cell layer with quiescent centre-like transcriptional identity exhibits the ability to regenerate organs. In the middle cell layer, WUSCHEL-RELATED HOMEOBOX5 (WOX5) directly interacts with PLETHORA1 and 2 to promote TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1 expression for endogenous auxin production. WOX5 also interacts with the B-type ARABIDOPSIS RESPONSE REGULATOR12 (ARR12) and represses A-type ARRs to break the negative feedback loop in cytokinin signalling. Overall, the promotion of auxin production and the enhancement of cytokinin sensitivity are both required for pluripotency acquisition in the middle cell layer of callus for organ regeneration.
Project description:Transcriptional profiling of age-related change of callus formation capability in Arabidopsis hypocotyls Organogenesis in vitro consists of many aspects such as phytohormone perception, dedifferentiation of differentiated cell to acquire organogenic competence, and re-entry of quiescent cells into cell cycle. In this study, we established an in vitro experimental system to study the age-dependent callus formation capacity in Arabidopsis. Interestingly, mature (35- to 38-day-old) hypocotyl explants exhibited better callus-forming potential than that of juvenile (7- to 10-day-old), determined by callus growth rates. To explore genome-wide expression changes underlying the phenomenon of age-dependent callus formation, a transcriptome-based analysis was performed. Gene expression profiling indicated that age-dependent callus formation capacity was associated with changes in phytohormone (auxins, cytokinins, abscisic acid, brassinosteroids and gibberellins) homeostasis, epigenetic mechanism and the cell cycle regulation. Besides, we identified two groups of genes involved in age-dependent callus formation capacity: (1) positive regulatory and (2) negative regulatory categories, i.e. genes that were significantly up- or down-regulated during callus formation derived from mature explants, respectively. One gene encoding DNA-binding protein (VARIANT IN METHYLATION 1, VIM1) belonging to the positive regulatory category was selected for functional analysis and assessment of age-dependent callus formation capacity. Indeed, vim1 reduced the efficiency of callus formation in mature explants, but not in juvenile. The result suggests that VIM1 plays an important role in regulating age-dependent callus formation capacity. Taken together, the investigation will help to better understand the molecular regulatory mechanism of age-dependent callus formation.
Project description:Transcriptional profiling of age-related change of callus formation capability in Arabidopsis hypocotyls Organogenesis in vitro consists of many aspects such as phytohormone perception, dedifferentiation of differentiated cell to acquire organogenic competence, and re-entry of quiescent cells into cell cycle. In this study, we established an in vitro experimental system to study the age-dependent callus formation capacity in Arabidopsis. Interestingly, mature (35- to 38-day-old) hypocotyl explants exhibited better callus-forming potential than that of juvenile (7- to 10-day-old), determined by callus growth rates. To explore genome-wide expression changes underlying the phenomenon of age-dependent callus formation, a transcriptome-based analysis was performed. Gene expression profiling indicated that age-dependent callus formation capacity was associated with changes in phytohormone (auxins, cytokinins, abscisic acid, brassinosteroids and gibberellins) homeostasis, epigenetic mechanism and the cell cycle regulation. Besides, we identified two groups of genes involved in age-dependent callus formation capacity: (1) positive regulatory and (2) negative regulatory categories, i.e. genes that were significantly up- or down-regulated during callus formation derived from mature explants, respectively. One gene encoding DNA-binding protein (VARIANT IN METHYLATION 1, VIM1) belonging to the positive regulatory category was selected for functional analysis and assessment of age-dependent callus formation capacity. Indeed, vim1 reduced the efficiency of callus formation in mature explants, but not in juvenile. The result suggests that VIM1 plays an important role in regulating age-dependent callus formation capacity. Taken together, the investigation will help to better understand the molecular regulatory mechanism of age-dependent callus formation. Comparison of young and mature Arabidopsis hypocotyls either with or without auxin treatment for 1 day
Project description:In plant tissue culture, callus forms from detached explants in response to a high-auxin-to-low-cytokinin ratio on callus-inducing medium. Callus is a group of pluripotent cells because it can regenerate either roots or shoots in response to a low level of auxin on root-inducing medium or a high-cytokinin-to-low-auxin ratio on shoot-inducing medium, respectively1. However, our knowledge of the mechanism of pluripotency acquisition during callus formation is limited. On the basis of analyses at the single-cell level, we show that the tissue structure of Arabidopsis thaliana callus on callus-inducing medium is similar to that of the root primordium or root apical meristem, and the middle cell layer with quiescent centre-like transcriptional identity exhibits the ability to regenerate organs. In the middle cell layer, WUSCHEL-RELATED HOMEOBOX5 (WOX5) directly interacts with PLETHORA1 and 2 to promote TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1 expression for endogenous auxin production. WOX5 also interacts with the B-type ARABIDOPSIS RESPONSE REGULATOR12 (ARR12) and represses A-type ARRs to break the negative feedback loop in cytokinin signalling. Overall, the promotion of auxin production and the enhancement of cytokinin sensitivity are both required for pluripotency acquisition in the middle cell layer of callus for organ regeneration.
Project description:Wounding is a primary trigger of organ regeneration but how wound stress reactivates cell proliferation and promotes cellular reprogramming remains elusive. In this study we combined the transcriptome analysis with quantitative hormonal analysis to investigate how wounding induces callus formation in Arabidopsis thaliana. Our time-course RNA-seq analysis revealed that wounding induces dynamic transcriptional changes that can be categorized into five clusters with distinct temporal patterns. Gene ontology analyses uncovered that wounding modifies the expression of hormone biosynthesis and response genes, and quantitative analysis of endogenous plant hormones revealed accumulation of cytokinin prior to callus formation. Mutants defective in cytokinin synthesis and signalling display reduced efficiency in callus formation, indicating that de novo synthesis of cytokinin has major contribution in wound-induced callus formation. We further demonstrate that type-A ARABIDOPSIS RESPONSE REGULATOR (ARR)-mediated cytokinin signalling regulates the expression of CYCLIN D3;1 (CYCD3;1) and mutations in CYCD3;1 and its homologs CYCD3;2-3 cause defects in callus formation. Our transcriptome data, in addition, showed that wounding activates multiple developmental regulators, and we found novel roles of ETHYLENE RESPONSE FACTOR 115 (ERF115) and PLETHORA3 (PLT3), PLT5, PLT7 in wound-induced callus formation. Together, this study provides novel mechanistic insights into how wounding reactivates cell proliferation during callus formation.
Project description:In plant tissue culture, callus forms from detached explants in response to a high-auxin-to-low-cytokinin ratio on callus-inducing medium. Callus is a group of pluripotent cells because it can regenerate either roots or shoots in response to a low level of auxin on root-inducing medium or a high-cytokinin-to-low-auxin ratio on shoot-inducing medium, respectively1. However, our knowledge of the mechanism of pluripotency acquisition during callus formation is limited. On the basis of analyses at the single-cell level, we show that the tissue structure of Arabidopsis thaliana callus on callus-inducing medium is similar to that of the root primordium or root apical meristem, and the middle cell layer with quiescent centre-like transcriptional identity exhibits the ability to regenerate organs. In the middle cell layer, WUSCHEL-RELATED HOMEOBOX5 (WOX5) directly interacts with PLETHORA1 and 2 to promote TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1 expression for endogenous auxin production. WOX5 also interacts with the B-type ARABIDOPSIS RESPONSE REGULATOR12 (ARR12) and represses A-type ARRs to break the negative feedback loop in cytokinin signalling. Overall, the promotion of auxin production and the enhancement of cytokinin sensitivity are both required for pluripotency acquisition in the middle cell layer of callus for organ regeneration.
Project description:Profiling the transcriptome of the early stage of Arabidopsis callus induction variable_1 = root explants variable_2 = aerial organ explants variable_3 = 0 h on callus inducing medium variable_4 = 12 h on callus inducing medium variable_5 = 24 h on callus inducing medium variable_6 = 48 h on callus inducing medium variable_7 = 96 h on callus inducing medium