Abstract
The Mediator complex controls transcription of most eukaryotic genes with individual subunits required for the control of particular gene regulons in response to various perturbations. In this study, we reveal the roles of the plant Mediator subunits MED16, MED14, and MED2 in regulating transcription in response to the phytohormone abscisic acid (ABA) and we determine which cis elements are under their control. Using synthetic promoter reporters we established an effective system for testing relationships between subunits and specific cis-acting motifs in protoplasts. Our results demonstrate that MED16, MED14, and MED2 are required for the full transcriptional activation by ABA of promoters containing both the ABRE (ABA-responsive element) and DRE (drought-responsive element). Using synthetic promoter motif concatamers, we showed that ABA-responsive activation of the ABRE but not the DRE motif was dependent on these three Mediator subunits. Furthermore, the three subunits were required for the control of water loss from leaves but played no role in ABA-dependent growth inhibition, highlighting specificity in their functions. Our results identify new roles for three Mediator subunits, provide a direct demonstration of their function and highlight that our experimental approach can be utilized to identify the function of subunits of plant transcriptional regulators.
Highlights
Transcriptional regulators are commonly assembled in multi-subunit protein complexes that regulate varied transcriptional responses
We showed that the Mediator complex subunit Mediator subunit 16 (MED16) is required for cold-responsive COR gene activation by the CBF (DREB1) transcription factor (TF), which bind the CRT/DRE motif (Yamaguchi-Shinozaki and Shinozaki, 1994)
To investigate the possibility that MED16 could control drought-responsive expression via the abscisic acid (ABA)-dependent pathway leading to activation of the ABRE motif, we quantified the expression of well-studied ABA- and drought-inducible genes, in med16 mutants following exogenous application of ABA
Summary
Transcriptional regulators are commonly assembled in multi-subunit protein complexes that regulate varied transcriptional responses. The Mediator complex is required for constitutive as well as inducible gene expression (Ansari et al, 2009) and consists of approximately 25–35 subunits, depending on the species (Bjorklund and Gustafsson, 2005). The head takes part in multiple associations with Pol II and is linked to the tail by the middle; the tail is considered to be the main site of transcription factor (TF) binding and as such, is the least conserved part of the complex (Maji et al, 2019), with tail subunits showing the most inter-species divergence (Bourbon, 2008; Conaway and Conaway, 2011). Individual subunits of the kinase module facilitate specific transcriptional events, for instance the expression of developmental and stress-responsive genes in plants (Gillmor et al, 2010; Ng et al, 2013). The homology of the complex subunits between species is weak at the sequence level the structural homology is remarkably well-conserved (Cai et al, 2009)
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