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Extensive modulation of the transcription factor transcriptome during somatic embryogenesis in Arabidopsis thaliana.

ABSTRACT: Molecular mechanisms controlling plant totipotency are largely unknown and studies on somatic embryogenesis (SE), the process through which already differentiated cells reverse their developmental program and become embryogenic, provide a unique means for deciphering molecular mechanisms controlling developmental plasticity of somatic cells. Among various factors essential for embryogenic transition of somatic cells transcription factors (TFs), crucial regulators of genetic programs, are believed to play a central role. Herein, we used quantitative real-time polymerase chain reaction (qRT-PCR) to identify TF genes affected during SE induced by in vitro culture in Arabidopsis thaliana. Expression profiles of 1,880 TFs were evaluated in the highly embryogenic Col-0 accession and the non-embryogenic tanmei/emb2757 mutant. Our study revealed 729 TFs whose expression changes during the 10-days incubation period of SE; 141 TFs displayed distinct differences in expression patterns in embryogenic versus non-embryogenic cultures. The embryo-induction stage of SE occurring during the first 5 days of culture was associated with a robust and dramatic change of the TF transcriptome characterized by the drastic up-regulation of the expression of a great majority (over 80%) of the TFs active during embryogenic culture. In contrast to SE induction, the advanced stage of embryo formation showed attenuation and stabilization of transcript levels of many TFs. In total, 519 of the SE-modulated TFs were functionally annotated and transcripts related with plant development, phytohormones and stress responses were found to be most abundant. The involvement of selected TFs in SE was verified using T-DNA insertion lines and a significantly reduced embryogenic response was found for the majority of them. This study provides comprehensive data focused on the expression of TF genes during SE and suggests directions for further research on functional genomics of SE.

SUBMITTER: Gliwicka M

PROVIDER: S-EPMC3714258 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Extensive modulation of the transcription factor transcriptome during somatic embryogenesis in Arabidopsis thaliana.

Gliwicka Marta M Nowak Katarzyna K Balazadeh Salma S Mueller-Roeber Bernd B Gaj Malgorzata D MD

PloS one 20130717 7

Molecular mechanisms controlling plant totipotency are largely unknown and studies on somatic embryogenesis (SE), the process through which already differentiated cells reverse their developmental program and become embryogenic, provide a unique means for deciphering molecular mechanisms controlling developmental plasticity of somatic cells. Among various factors essential for embryogenic transition of somatic cells transcription factors (TFs), crucial regulators of genetic programs, are believe ...[more]

PMID: 23874927

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Project description:Somatic embryogenesis (SE) is a process by which an embryo is derived from somatic tissue. Transcription factors (TFs) have been identified that control this process. One such TF that promotes SE is AGAMOUS-like 15 (AGL15). Prior work has shown that AGL15 can both induce and repress gene expression. One way this type of dual function TF works is via protein interactions, so a yeast 2-hybrid (Y2H) screen was undertaken. One intriguing protein with which AGL15 interacted in Y2H was LBD40. LBD40 encodes a LATERAL ORGAN BOUNDARIES (LOB)-domain TF that is unique to plants and is primarily expressed during seed development. Here, we confirm the AGL15-LBD40 interaction by quantitative assays and in planta co-immunoprecipation. We also document a role for LBD40, and the closely related protein LBD41, in supporting SE. To determine downstream genes potentially controlled by LBD40, chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) was used. More than 400 binding regions for LBD40 were consistently found genome-wide. To determine genes responsive to LBD40/41 accumulation, RNA-seq analysis of transcriptomes of wild-type control and loss-of-function lbd40/lbd41 was performed. Combining these datasets provides insight into genes directly and indirectly controlled by these LOB domain TFs. The gene ontology (GO) enrichment analysis of these regulated genes showed an overrepresentation of biological processes that are associated with SE, further indicating the importance of LBD40 in SE. This work provides insight into SE, a poorly understood, but essential process to generate transgenic plants to meet agricultural demands or test gene function. This manuscript reports on experiments to understand the role that LDB40, a TF, plays in support of SE by investigating genes directly and indirectly controlled by LBD40 and examining physical and genetic interactions with other TFs active in SE. We uncover targets of LBD40 and an interacting TF of the MADS family and investigate targets involvement in SE.

Project description:BackgroundSomatic embryogenesis (SE) exemplifies the unique developmental plasticity of plant cells. The regulatory processes, including epigenetic modifications controlling embryogenic reprogramming of cell transcriptome, have just started to be revealed.ResultsTo identify the genes of histone acetylation-regulated expression in SE, we analyzed global transcriptomes of Arabidopsis explants undergoing embryogenic induction in response to treatment with histone deacetylase inhibitor, trichostatin A (TSA). The TSA-induced and auxin (2,4-dichlorophenoxyacetic acid; 2,4-D)-induced transcriptomes were compared. RNA-seq results revealed the similarities of the TSA- and auxin-induced transcriptomic responses that involve extensive deregulation, mostly repression, of the majority of genes. Within the differentially expressed genes (DEGs), we identified the master regulators (transcription factors - TFs) of SE, genes involved in biosynthesis, signaling, and polar transport of auxin and NITRILASE-encoding genes of the function in indole-3-acetic acid (IAA) biosynthesis. TSA-upregulated TF genes of essential functions in auxin-induced SE, included LEC1/LEC2, FUS3, AGL15, MYB118, PHB, PHV, PLTs, and WUS/WOXs. The TSA-induced transcriptome revealed also extensive upregulation of stress-related genes, including those related to stress hormone biosynthesis. In line with transcriptomic data, TSA-induced explants accumulated salicylic acid (SA) and abscisic acid (ABA), suggesting the role of histone acetylation (Hac) in regulating stress hormone-related responses during SE induction. Since mostly the adaxial side of cotyledon explant contributes to SE induction, we also identified organ polarity-related genes responding to TSA treatment, including AIL7/PLT7, RGE1, LBD18, 40, HB32, CBF1, and ULT2. Analysis of the relevant mutants supported the role of polarity-related genes in SE induction.ConclusionThe study results provide a step forward in deciphering the epigenetic network controlling embryogenic transition in somatic cells of plants.

Project description:Several genes encoding transcription factors (TFs) were indicated to have a key role in the induction of somatic embryogenesis (SE), which is triggered in the somatic cells of plants. In order to further explore the genetic regulatory network that is involved in the embryogenic transition induced in plant somatic cells, micro-RNA (miRNAs) molecules, the products of MIRNA (MIR) genes and the common regulators of TF transcripts, were analyzed in an embryogenic culture of Arabidopsis thaliana. In total, the expression of 190 genes of the 114 MIRNA families was monitored during SE induction and the levels of the primary (pri-miRNAs) transcripts vs. the mature miRNAs were investigated. The results revealed that the majority (98%) of the MIR genes were active and that most of them (64%) were differentially expressed during SE. A distinct attribute of the MIR expression in SE was the strong repression of MIR transcripts at the early stage of SE followed by their significant up-regulation in the advanced stage of SE. Comparison of the mature miRNAs vs. pri-miRNAs suggested that the extensive post-transcriptional regulation of miRNA is associated with SE induction. Candidate miRNA molecules of the assumed function in the embryogenic response were identified among the mature miRNAs that had a differential expression in SE, including miR156, miR157, miR159, miR160, miR164, miR166, miR169, miR319, miR390, miR393, miR396, and miR398. Consistent with the central role of phytohormones and stress factors in SE induction, the functions of the candidate miRNAs were annotated to phytohormone and stress responses. To confirm the functions of the candidate miRNAs in SE, the expression patterns of the mature miRNAs and their presumed targets were compared and regulatory relation during SE was indicated for most of the analyzed miRNA-target pairs. The results of the study contribute to the refinement of the miRNA-controlled regulatory pathways that operate during embryogenic induction in plants and provide a valuable platform for the identification of the genes that are targeted by the candidate miRNAs in SE induction.

Project description:BackgroundSeed germination involves progression from complete metabolic dormancy to a highly active, growing seedling. Many factors regulate germination and these interact extensively, forming a complex network of inputs that control the seed-to-seedling transition. Our understanding of the direct regulation of gene expression and the dynamic changes in the epigenome and small RNAs during germination is limited. The interactions between genome, transcriptome and epigenome must be revealed in order to identify the regulatory mechanisms that control seed germination.ResultsWe present an integrated analysis of high-resolution RNA sequencing, small RNA sequencing and MethylC sequencing over ten developmental time points in Arabidopsis thaliana seeds, finding extensive transcriptomic and epigenomic transformations associated with seed germination. We identify previously unannotated loci from which messenger RNAs are expressed transiently during germination and find widespread alternative splicing and divergent isoform abundance of genes involved in RNA processing and splicing. We generate the first dynamic transcription factor network model of germination, identifying known and novel regulatory factors. Expression of both microRNA and short interfering RNA loci changes significantly during germination, particularly between the seed and the post-germinative seedling. These are associated with changes in gene expression and large-scale demethylation observed towards the end of germination, as the epigenome transitions from an embryo-like to a vegetative seedling state.ConclusionsThis study reveals the complex dynamics and interactions of the transcriptome and epigenome during seed germination, including the extensive remodelling of the seed DNA methylome from an embryo-like to vegetative-like state during the seed-to-seedling transition. Data are available for exploration in a user-friendly browser at https://jbrowse.latrobe.edu.au/germination_epigenome .

Dataset Information

Extensive modulation of the transcription factor transcriptome during somatic embryogenesis in Arabidopsis thaliana.

Publications

Extensive modulation of the transcription factor transcriptome during somatic embryogenesis in Arabidopsis thaliana.

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