REVEILLE1, a Myb-like transcription factor, integrates the circadian clock and auxin pathways.

The circadian clock modulates expression of a large fraction of the Arabidopsis genome and affects many aspects of plant growth and development. We have discovered one way in which the circadian system regulates hormone signaling, identifying a node that links the clock and auxin networks. Auxin plays key roles in development and responses to environmental cues, in part through regulation of plant growth. We have characterized REVEILLE1 (RVE1), a Myb-like, clock-regulated transcription factor that is homologous to the central clock genes CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY). Despite this homology, inactivation of RVE1 does not affect circadian rhythmicity but instead causes a growth phenotype, indicating this factor is a clock output affecting plant development. CCA1 regulates growth via the bHLH transcription factors PHYTOCHROME INTERACTING FACTOR4 (PIF4) and PIF5, but RVE1 acts independently of these genes. RVE1 instead controls auxin levels, promoting free auxin production during the day but having no effect during the night. RVE1 positively regulates the expression of the auxin biosynthetic gene YUCCA8 (YUC8), providing a mechanism for its growth-promoting effects. RVE1 is therefore a node that connects two important signaling networks that coordinate plant growth with rhythmic changes in the environment.

The FLX gene of Arabidopsis is required for FRI-dependent activation of FLC expression.

The Arabidopsis FLOWERING LOCUS C (FLC) gene encodes a MADS box protein that acts as a dose-dependent repressor of flowering. Mutants and ecotypes with elevated expression of FLC are late flowering and vernalization responsive. In this study we describe an early flowering mutant in the C24 ecotype, flc expressor (flx), that has reduced expression of FLC. FLX encodes a protein of unknown function with putative leucine zipper domains. FLX is required for FRIGIDA (FRI)-mediated activation of FLC but not for activation of FLC in autonomous pathway mutants. FLX is also required for expression of the FLC paralogs MADS AFFECTING FLOWERING 1 (MAF1) and MAF2.

Differential gene expression and subcellular targeting of Arabidopsis glutathione S-transferase F8 is achieved through alternative transcription start sites.

Glutathione S-transferases (GSTs) play major roles in the protection of plants from biotic and abiotic stresses through the detoxification of xenobiotics and toxic endogenous products. This report describes additional complexity in the regulation of the well characterized stress-responsive Arabidopsis thaliana GSTF8 promoter. This complexity results from the use of multiple transcription start sites (TSS) to give rise to alternate GSTF8 transcripts with the potential to produce two in-frame proteins differing only in their N-terminal sequence. In addition to the originally mapped TSS (Chen, W., Chao, G., and Singh, K. B. (1996) Plant J. 10, 955-966), a further nine TSS have been identified, with the majority clustered into a distinct group. The most 3' TSS gives rise to the major message (GSTF8-S) and the shorter form of the protein, whereas those originating from upstream TSS (GSTF8-L) are more weakly expressed and encode for the larger form of the protein. Differential tissue-specific and stress-responsive expression patterns were observed (e.g. GSTF8-L is more highly expressed in leaves compared with roots, whereas GSTF8-S expression has the opposite pattern and is much more stress-responsive). Analysis of GSTF8-L and GSTF8-S proteins demonstrated that GSTF8-L is solely targeted to plastids, whereas GSTF8-S is cytoplasmic. In silico analysis revealed potential conservation of GSTF8-S across a wide range of plants; in contrast, conservation of GSTF8-L was confined to the Brassicaceae. These studies demonstrate that alternate TSS of the GSTF8 promoter are used to confer differential tissue-specific and stress-responsive expression patterns as well as to target the same protein to two different subcellular localizations.

PsfR, a factor that stimulates psbAI expression in the cyanobacterium Synechococcus elongatus PCC 7942.

In this paper a gene (psfR) is reported that regulates psbAI activity in Synechococcus elongatus, a unicellular photoautotrophic cyanobacterium that carries out oxygenic (plant-type) photosynthesis and exhibits global circadian regulation of gene expression. In S. elongatus, a family of three psbA genes encodes the D1 protein of the photosystem II reaction centre. Overexpression of psfR results in increased expression of psbAI, but does not affect the circadian timing of psbAI expression. psfR overexpression affected some, but not all of the genes routinely surveyed for circadian expression. PsfR acts (directly or indirectly) on the psbAI basal promoter region. psfR knockout mutants exhibit wild-type psbAI expression, suggesting that other factors can regulate psbAI expression in the absence of functional PsfR. PsfR contains two receiver-like domains (found in bacterial two-component signal transduction systems), one of which lacks the conserved aspartyl residue required for phosphoryl transfer. PsfR also contains a GGDEF domain. The presence of these domains and the absence of a detectable conserved DNA-binding domain suggest that PsfR may regulate psbAI expression via protein-protein interactions or GGDEF activity (the production of cyclic dinucleotides) rather than direct interaction with the psbAI promoter.

HY5, Circadian Clock-Associated 1, and a cis-element, DET1 dark response element, mediate DET1 regulation of chlorophyll a/b-binding protein 2 expression.

DET1 is a pleiotropic regulator of Arabidopsis development and controls the expression of many light-regulated genes. To gain a better understanding of the mechanism by which DET1 controls transcription from light-regulated promoters, we identified elements in the chlorophyll a/b-binding protein 2 (CAB2) promoter that are required for DET1-mediated expression. Using a series of reporter constructs in which the luciferase gene is controlled by CAB2 promoter fragments, we defined two DET1-responsive elements in the CAB2 promoter that are essential for proper CAB2 transcription. A 40-bp DET1 dark-response element (DtRE) is required for both dark and root-specific repression of CAB2, whereas the known CAB upstream factor-1 element is required for DET1 activation-associated effects in the light and repression in the roots. HY5, a factor that binds CAB upstream factor-1, is also required for DET1 effects in the light. DtRE binds two distinct activities in Arabidopsis seedling extracts: a novel activity with binding site CAAAACGC that we have named CAB2 DET1-associated factor 1 plus an activity that is likely to be the myb transcription factor Circadian Clock-Associated 1. Both activities are altered in dark-grown det1 extracts as compared with wild type, correlating a change in extractable DNA binding activity with a major change in CAB2 expression. We conclude that DET1 represses the CAB2 promoter in the dark by regulating the binding of two factors, CAB2 DET1-associated factor 1 and Circadian Clock-Associated 1, to the DtRE.