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Abstract Transcription factors carry long intrinsically disordered regions often containing multiple activation domains. Despite numerous recent high‐throughput identifications and characterizations of activation domains, the interplay between sequence motifs, activation domains, and regulator binding in intrinsically disordered transcription factor regions remains unresolved. Here, we map sequence motifs and activation domains in anArabidopsis thalianaNAC transcription factor clade, revealing that although sequence motifs and activation domains often coincide, no systematic overlap exists. Biophysical analyses using NMR spectroscopy show that the long intrinsically disordered region of senescence‐associated transcription factor ANAC046 is devoid of residual structure. We identify two activation domain/sequence motif regions, one at each end that both bind a panel of six positive and negative regulator domains from biologically relevant regulators promiscuously. Binding affinities measured using isothermal titration calorimetry reveal a hierarchy for regulator binding of the two ANAC046 activation domain/sequence motif regions defining these as regulatory hotspots. Despite extensive dynamic intramolecular contacts along the disordered chain revealed using paramagnetic relaxation enhancement experiments and simulations, the regions remain uncoupled in binding. Together, the results imply rheostatic regulation by ANAC046 through concentration‐dependent regulator competition, a mechanism likely mirrored in other transcription factors with distantly located activation domains. 

Apical hook opening is crucial for seedling establishment and is regulated by unequal distribution of the hormone auxin through unknown mechanisms. In this issue of Developmental Cell, Walia et al. 4 demonstrate that apical hook opening is an output of tissue-wide forces; auxin and cell wall integrity (CWI) signaling interact to restrict elongation to the concave side of the apical hook. 

Abstract TheArabidopsis thalianaDREB2A transcription factor interacts with the negative regulator RCD1 and the ACID domain of subunit 25 of the transcriptional co-regulator mediator (Med25) to integrate stress signals for gene expression, with elusive molecular interplay. Using biophysical and structural analyses together with high-throughput screening, we reveal a bivalent binding switch in DREB2A containing an ACID-binding motif (ABS) and the known RCD1-binding motif (RIM). The RIM is lacking in a stress-induced DREB2A splice variant with retained transcriptional activity. ABS and RIM bind to separate sites on Med25-ACID, and NMR analyses show a structurally heterogeneous complex deriving from a DREB2A-ABS proline residue populatingcis- andtrans-isomers with remote impact on the RIM. Thecis-isomer stabilizes an α-helix, while thetrans-isomer may introduce energetic frustration facilitating rapid exchange between activators and repressors. Thus, DREB2A uses a post-transcriptionally and post-translationally modulated switch for transcriptional regulation. 

Gene expression in Arabidopsis is regulated by more than 1,900 transcription factors (TFs), which have been identified genome-wide by the presence of well-conserved DNA-binding domains. Activator TFs contain activation domains (ADs) that recruit coactivator complexes; however, for nearly all Arabidopsis TFs, we lack knowledge about the presence, location and transcriptional strength of their ADs1. To address this gap, here we use a yeast library approach to experimentally identify Arabidopsis ADs on a proteome-wide scale, and find that more than half of the Arabidopsis TFs contain an AD. We annotate 1,553 ADs, the vast majority of which are, to our knowledge, previously unknown. Using the dataset generated, we develop a neural network to accurately predict ADs and to identify sequence features that are necessary to recruit coactivator complexes. We uncover six distinct combinations of sequence features that result in activation activity, providing a framework to interrogate the subfunctionalization of ADs. Furthermore, we identify ADs in the ancient AUXIN RESPONSE FACTOR family of TFs, revealing that AD positioning is conserved in distinct clades. Our findings provide a deep resource for understanding transcriptional activation, a framework for examining function in intrinsically disordered regions and a predictive model of ADs. 

The phytohormone auxin regulates nearly every aspect of plant development. Transcriptional responses to auxin are driven by the activities of the AUXIN RESPONSE FACTOR family of transcription factors. ARF19 (AT1G19220) is critical in the auxin signaling pathway and has previously been shown to undergo protein condensation to tune auxin responses in the root. However, ARF19 condensation dynamics in other organs has not yet been described. In the Arabidopsis stomatal lineage, we found that ARF19 cytoplasmic condensates are enriched in guard cells and pavement cells, terminally differentiated cells in the leaf epidermis. This result is consistent with previous studies showing ARF19 condensation in mature root tissues. Our data reveal that the sequestration of ARF19 into cytoplasmic condensation in differentiated leaf epidermal cells is similar to root-specific condensation patterns. 

Abstract This review highlights recent literature on biomolecular condensates in plant development and discusses challenges for fully dissecting their functional roles. Plant developmental biology has been inundated with descriptive examples of biomolecular condensate formation, but it is only recently that mechanistic understanding has been forthcoming. Here, we discuss recent examples of potential roles biomolecular condensates play at different stages of the plant life cycle. We group these examples based on putative molecular functions, including sequestering interacting components, enhancing dwell time, and interacting with cytoplasmic biophysical properties in response to environmental change. We explore how these mechanisms could modulate plant development in response to environmental inputs and discuss challenges and opportunities for further research into deciphering molecular mechanisms to better understand the diverse roles that biomolecular condensates exert on life. 

Abstract Auxin critically regulates plant growth and development. Auxin-driven transcriptional responses are mediated through the AUXIN RESPONSE FACTOR (ARF) family of transcription factors. ARF protein condensation attenuates ARF activity, resulting in dramatic shifts in the auxin transcriptional landscape. Here, we perform a forward genetics screen for ARF hypercondensation, identifying an F-box protein, which we named AUXIN RESPONSE FACTOR F-BOX1 (AFF1). Functional characterization of SCF^AFF1revealed that this E3 ubiquitin ligase directly interacts with ARF19 and ARF7 to regulate their accumulation, condensation, and nucleo-cytoplasmic partitioning. Mutants defective inAFF1display attenuated auxin responsiveness, and developmental defects, suggesting that SCF^AFF1-mediated regulation of ARF protein drives aspects of auxin response and plant development.