Search for: All records

AGAMOUS-like 15 (AGL15) is a member of the MADS-domain transcription factor (TF) family. MADS proteins are named for a conserved domain that was originally from an acronym derived from genes expressed in a variety of eukaryotes (MCM1-AGAMOUS-DEFICIENS-SERUM RESPONSE FACTOR). In plants, this family has expanded greatly, with more than one-hundred members generally found in dicots, and the proteins encoded by these genes have often been associated with developmental identity. AGL15 transcript and protein accumulate primarily in embryos and has been found to promote an important process called plant regeneration via somatic embryogenesis (SE). To understand how this TF performs this function, we have previously used microarray technologies to assess direct and indirect responsive targets of this TF. We have now revisited this question using next generation sequencing (NGS) to both characterize in vivo binding sites for AGL15 as well as response to the accumulation of AGL15. We compared these data to the prior microarray results to evaluate the different platforms. The new NGS data brought to light an interaction with brassinosteroid (BR) hormone signaling that was “missed” in prior Gene Ontology analysis from the microarray studies. 

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 andin plantaco‐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‐functionlbd40/lbd41was 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.

 

Plants have amazing regenerative properties with single somatic cells, or groups of cells able to give rise to fully formed plants. One means of regeneration is somatic embryogenesis, by which an embryonic structure is formed that “converts” into a plantlet. Somatic embryogenesis has been used as a model for zygotic processes that are buried within layers of maternal tissues. Understanding mechanisms of somatic embryo induction and development are important as a more accessible model for seed development. We rely on seed development not only for most of our caloric intake, but also as a delivery system for engineered crops to meet agricultural challenges. Regeneration of transformed cells is needed for this applied work as well as basic research to understand gene function. Here we focus on a MADS-domain transcription factor, AGAMOUS-Like15 (AGL15) that shows a positive correlation between accumulation levels and capacity for somatic embryogenesis. We relate AGL15 function to other transcription factors, hormones, and epigenetic modifiers involved in somatic embryo development. 

In contrast to animals, adult organs in plants are not formed during embryogenesis but generated from meristematic cells as plants advance through development. Plant development involves a succession of different phenotypic stages and the transition between these stages is termed phase transition. Phase transitions need to be tightly regulated and coordinated to ensure they occur under optimal seasonal, environmental conditions. Polycarpic perennials transition through vegetative stages and the mature, reproductive stage many times during their lifecycles and, in both perennial and annual species, environmental factors and culturing methods can reverse the otherwise unidirectional vector of plant development. Epigenetic factors regulating gene expression in response to internal cues and external (environmental) stimuli influencing the plant’s phenotype and development have been shown to control phase transitions. How developmental and environmental cues interact to epigenetically alter gene expression and influence these transitions is not well understood, and understanding this interaction is important considering the current climate change scenarios, since epigenetic maladaptation could have catastrophic consequences for perennial plants in natural and agricultural ecosystems. Here, we review studies focusing on the epigenetic regulators of the vegetative phase change and highlight how these mechanisms might act in exogenously induced plant rejuvenation and regrowth following stress. 

AGAMOUS-like 15 (AGL15) is a member of the MADS domain family of transcription factors (TFs) that can directly induce and repress target gene expression, and for which promotion of somatic embryogenesis (SE) is positively correlated with accumulation. An ethylene-responsive element binding factor-associated amphiphilic repression (EAR) motif of form LxLxL within the carboxyl-terminal domain of AGL15 was shown to be involved in repression of gene expression. Here, we examine whether AGL15′s ability to repress gene expression is needed to promote SE. While a form of AGL15 where the LxLxL is changed to AxAxA can still promote SE, another form with a strong transcriptional activator at the carboxy-terminal end, does not promote SE and, in fact, is detrimental to SE development. Select target genes were examined for response to the different forms of AGL15. 

Seeds are essential for human civilization, so understanding the molecular events underpinning seed development and the zygotic embryo it contains is important. In addition, the approach of somatic embryogenesis is a critical propagation and regeneration strategy to increase desirable genotypes, to develop new genetically modified plants to meet agricultural challenges, and at a basic science level, to test gene function. We briefly review some of the transcription factors (TFs) involved in establishing primary and apical meristems during zygotic embryogenesis, as well as TFs necessary and/or sufficient to drive somatic embryo programs. We focus on the model plant Arabidopsis for which many tools are available, and review as well as speculate about comparisons and contrasts between zygotic and somatic embryo processes. 

AGAMOUS-Like 18 (AGL18) is a MADS domain transcription factor (TF) that is structurally related to AGL15. Here we show that, like AGL15, AGL18 can promote somatic embryogenesis (SE) when ectopically expressed in Arabidopsis (Arabidopsis thaliana). Based on loss-of-function mutants, AGL15 and AGL18 have redundant functions in developmental processes such as SE. To understand the nature of this redundancy, we undertook a number of studies to look at the interaction between these factors. We studied the genome-wide direct targets of AGL18 to characterize its roles at the molecular level using chromatin immunoprecipitation (ChIP)-SEQ combined with RNA-SEQ. The results demonstrated that AGL18 binds to thousands of sites in the genome. Comparison of ChIP-SEQ data for AGL15 and AGL18 revealed substantial numbers of genes bound by both AGL15 and AGL18, but there were also differences. Gene ontology analysis revealed that target genes were enriched for seed, embryo, and reproductive development as well as hormone and stress responses. The results also demonstrated that AGL15 and AGL18 interact in a complex regulatory loop, where AGL15 inhibited transcript accumulation of AGL18, while AGL18 increased AGL15 transcript accumulation. Co-immunoprecipitation revealed an interaction between AGL18 and AGL15 in somatic embryo tissue. The binding and expression analyses revealed a complex crosstalk and interactions among embryo TFs and their target genes. In addition, our study also revealed that phosphorylation of AGL18 and AGL15 was crucial for the promotion of SE.

 

Arabidopsis thalianaABSCISIC ACID INSENSITIVE3 (ABI3) is a transcription factor in the B3 domain family. ABI3, along with B3 domain transcription factors LEAFY COTYLEDON2 (LEC2) and FUSCA3 (FUS3), and LEC1, a subunit of the CCAAT box‐binding complex, form the so‐called LAFL network to control various aspects of seed development and maturation. ABI3 also contributes to the abscisic acid (ABA) response. We report on chromatin immunoprecipitation‐tiling array experiments to map binding sites for ABI3 globally. We also assessed transcriptomes in response to ABI3 by comparing developingabi3‐5and wild‐type seeds and combined this information to ascertain direct and indirect responsive ABI3 target genes. ABI3 can induce and repress its transcription of target genes directly and some intriguing differences exist incismotifs between these groups of genes. Directly regulated targets reflect the role of ABI3 in seed maturation, desiccation tolerance, entry into a quiescent state and longevity. Interestingly, ABI3 directly represses a gene encoding a microRNA (MIR160B) that targetsAUXIN RESPONSE FACTOR(ARF)10andARF16that are involved in establishment of dormancy. In addition, ABI3, like FUS3, regulates genes encodingMIR156but while FUS3 only induces genes encoding this product, ABI3 induces these genes during the early stages of seed development, but represses these genes during late development. The interplay between ABI3, the otherLAFLgenes, and theVP1/ABI3‐LIKE(VAL) genes, which are involved in the transition to seedling development are examined and reveal complex interactions controlling development.