A function of TPL/TBL1-type corepressors is to nucleate the assembly of the preinitiation complex.

The plant corepressor TPL is recruited to diverse chromatin contexts, yet its mechanism of repression remains unclear. Previously, we leveraged the fact that TPL retains its function in a synthetic transcriptional circuit in the yeast model Saccharomyces cerevisiae to localize repressive function to two distinct domains. Here, we employed two unbiased whole-genome approaches to map the physical and genetic interactions of TPL at a repressed locus.

A history-dependent integrase recorder of plant gene expression with single-cell resolution.

During development, most cells experience a progressive restriction of fate that ultimately results in a fully differentiated mature state. Understanding more about the gene expression patterns that underlie developmental programs can inform engineering efforts for new or optimized forms. Here, we present a four-state integrase-based recorder of gene expression history and demonstrate its use in tracking gene expression events in Arabidopsis thaliana in two developmental contexts: lateral root initiation and stomatal differentiation.

A conserved function of corepressors is to nucleate assembly of the transcriptional preinitiation complex.

The plant corepressor TPL is recruited to diverse chromatin contexts, yet its mechanism of repression remains unclear. Previously, we have leveraged the fact that TPL retains its function in a synthetic transcriptional circuit in the yeast model to localize repressive function to two distinct domains. Here, we employed two unbiased whole genome approaches to map the physical and genetic interactions of TPL at a repressed locus.

Building a pipeline to identify and engineer constitutive and repressible promoters.

To support the increasingly complex circuits needed for plant synthetic biology applications, additional constitutive promoters are essential. Reusing promoter parts can lead to difficulty in cloning, increased heterogeneity between transformants, transgene silencing and trait instability. We have developed a pipeline to identify genes that have stable expression across a wide range of tissues at different developmental stages and have identified a number of promoters that are well expressed in both transient () and stable () transformation assays.

A comparative analysis of stably expressed genes across diverse angiosperms exposes flexibility in underlying promoter architecture.

Promoters regulate both the amplitude and pattern of gene expression-key factors needed for optimization of many synthetic biology applications. Previous work in Arabidopsis found that promoters that contain a TATA-box element tend to be expressed only under specific conditions or in particular tissues, while promoters that lack any known promoter elements, thus designated as Coreless, tend to be expressed more uniformly. To test whether this trend represents a conserved promoter design rule, we identified stably expressed genes across multiple angiosperm species using publicly available RNA-seq data.

A comparative analysis of stably expressed genes across diverse angiosperms exposes flexibility in underlying promoter architecture.

Promoters regulate both the amplitude and pattern of gene expression-key factors needed for optimization of many synthetic biology applications. Previous work in found that promoters that contain a TATA-box element tend to be expressed only under specific conditions or in particular tissues, while promoters which lack any known promoter elements, thus designated as Coreless, tend to be expressed more ubiquitously. To test whether this trend represents a conserved promoter design rule, we identified stably expressed genes across multiple angiosperm species using publicly available RNA-seq data.

An integrase toolbox to record gene-expression during plant development.

There are many open questions about the mechanisms that coordinate the dynamic, multicellular behaviors required for organogenesis. Synthetic circuits that can record in vivo signaling networks have been critical in elucidating animal development. Here, we report on the transfer of this technology to plants using orthogonal serine integrases to mediate site-specific and irreversible DNA recombination visualized by switching between fluorescent reporters.

A single helix repression domain is functional across diverse eukaryotes.

The corepressor TOPLESS (TPL) and its paralogs coordinately regulate a large number of genes critical to plant development and immunity. As in many members of the larger pan-eukaryotic Tup1/TLE/Groucho corepressor family, TPL contains a Lis1 Homology domain (LisH), whose function is not well understood. We have previously found that the LisH in TPL-and specifically the N-terminal 18 amino acid alpha-helical region (TPL-H1)-can act as an autonomous repression domain.

Decoding and recoding plant development.

The development of multicellular organisms has been studied for centuries, yet many critical events and mechanisms of regulation remain challenging to observe directly. Early research focused on detailed observational and comparative studies. Molecular biology has generated insights into regulatory mechanisms, but only for a limited number of species.

The Systems and Synthetic Biology of Auxin.

Auxin biology as a field has been at the forefront of advances in delineating the structures, dynamics, and control of plant growth networks. Advances have been enabled by combining the complementary fields of top-down, holistic systems biology and bottom-up, build-to-understand synthetic biology. Continued collaboration between these approaches will facilitate our understanding of and ability to engineer auxin's control of plant growth, development, and physiology.

Repression by the TOPLESS corepressor requires association with the core mediator complex.

The plant corepressor TOPLESS (TPL) is recruited to a large number of loci that are selectively induced in response to developmental or environmental cues, yet the mechanisms by which it inhibits expression in the absence of these stimuli are poorly understood. Previously, we had used the N-terminus of TPL to enable repression of a synthetic auxin response circuit in (yeast). Here, we leveraged the yeast system to interrogate the relationship between TPL structure and function, specifically scanning for repression domains.

A single-cell view of the transcriptome during lateral root initiation in Arabidopsis thaliana.

Root architecture is a major determinant of plant fitness and is under constant modification in response to favorable and unfavorable environmental stimuli. Beyond impacts on the primary root, the environment can alter the position, spacing, density, and length of secondary or lateral roots. Lateral root development is among the best-studied examples of plant organogenesis, yet there are still many unanswered questions about its earliest steps.

Expansion and innovation in auxin signaling: where do we grow from here?

The phytohormone auxin plays a role in almost all growth and developmental responses. The primary mechanism of auxin action involves the regulation of transcription via a core signaling pathway comprising proteins belonging to three classes: receptors, co-receptor/co-repressors and transcription factors. Recent studies have revealed that auxin signaling can be traced back at least as far as the transition to land.

It's Morphin' time: how multiple signals converge on ARF transcription factors to direct development.

Plant development programs are constantly updated by information about environmental conditions, currently available resources, and sites of active organogenesis. Much of this information is encoded in modifications of transcription factors that lead to changes in their relative abundance, activity and localization. Recent work on the Auxin Response Factor family of transcription factors has highlighted the large diversity of such modifications, as well as how they may work synergistically or antagonistically to regulate downstream responses.

A Synthetic Approach Allows Rapid Characterization of the Maize Nuclear Auxin Response Circuit.

Auxin plays a key role across all land plants in growth and developmental processes. Although auxin signaling function has diverged and expanded, differences in the molecular functions of signaling components have largely been characterized in Arabidopsis (). Here, we used the nuclear Auxin Response Circuit recapitulated in yeast () system to functionally annotate maize () auxin signaling components, focusing on genes expressed during the development of ear and tassel inflorescences.

Engineering Synthetic Signaling in Plants.

Synthetic signaling is a branch of synthetic biology that aims to understand native genetic regulatory mechanisms and to use these insights to engineer interventions and devices that achieve specified design parameters. Applying synthetic signaling approaches to plants offers the promise of mitigating the worst effects of climate change and providing a means to engineer crops for entirely novel environments, such as those in space travel. The ability to engineer new traits using synthetic signaling methods will require standardized libraries of biological parts and methods to assemble them; the decoupling of complex processes into simpler subsystems; and mathematical models that can accelerate the design-build-test-learn cycle.

Specificity in Auxin Responses Is Not Explained by the Promoter Preferences of Activator ARFs.

The plant growth hormone auxin regulates development via a family of transcription factors that share promoter sequence preferences, despite activating different genetic networks.

Mapping and Dynamics of Regulatory DNA in Maturing Siliques.

The genome is reprogrammed during development to produce diverse cell types, largely through altered expression and activity of key transcription factors. The accessibility and critical functions of epidermal cells have made them a model for connecting transcriptional events to development in a range of model systems. In and many other plants, fertilization triggers differentiation of specialized epidermal seed coat cells that have a unique morphology caused by large extracellular deposits of polysaccharides.

Auxin promotion of seedling growth via ARF5 is dependent on the brassinosteroid-regulated transcription factors BES1 and BEH4.

Seedlings must continually calibrate their growth in response to the environment. Auxin and brassinosteroids (BRs) are plant hormones that work together to control growth responses during photomorphogenesis. We used our previous analysis of promoter architecture in an auxin and BR target gene to guide our investigation into the broader molecular bases and biological relevance of transcriptional co-regulation by these hormones.

Accelerating structure-function mapping using the ViVa webtool to mine natural variation.

Thousands of sequenced genomes are now publicly available capturing a significant amount of natural variation within plant species; yet, much of these data remain inaccessible to researchers without significant bioinformatics experience. Here, we present a webtool called ViVa (Visualizing Variation) which aims to empower any researcher to take advantage of the amazing genetic resource collected in the 1001 Genomes Project (http://1001genomes.org).

Synthetic Bistability and Differentiation in Yeast.

Engineered systems that control cellular differentiation and pattern formation are essential for applications like tissue engineering, biomaterial fabrication, and synthetic ecosystems. Synthetic circuits that can take on multiple states have been made to engineer multicellular systems. However, how to use these states to drive interesting cellular behavior remains challenging.

The DNA binding landscape of the maize AUXIN RESPONSE FACTOR family.

AUXIN RESPONSE FACTORS (ARFs) are plant-specific transcription factors (TFs) that couple perception of the hormone auxin to gene expression programs essential to all land plants. As with many large TF families, a key question is whether individual members determine developmental specificity by binding distinct target genes. We use DAP-seq to generate genome-wide in vitro TF:DNA interaction maps for fourteen maize ARFs from the evolutionarily conserved A and B clades.

Synthetic hormone-responsive transcription factors can monitor and re-program plant development.

Developmental programs sculpt plant morphology to meet environmental challenges, and these same programs have been manipulated to increase agricultural productivity (Doebley et al., 1997; Khush, 2001). Hormones coordinate these programs, creating chemical circuitry (Vanstraelen and Benková, 2012) that has been represented in mathematical models (Refahi et al.

Phytochrome B regulates resource allocation in Brassica rapa.

Crop biomass and yield are tightly linked to how the light signaling network translates information about the environment into allocation of resources, including photosynthates. Once activated, the phytochrome (phy) class of photoreceptors signal and re-deploy carbon resources to alter growth, plant architecture, and reproductive timing. Most of the previous characterization of the light-modulated growth program has been performed in the reference plant Arabidopsis thaliana.