Transcriptional Regulation in Plants: The Importance of Combinatorial Control

Combinatorial control: use of a discrete number of transcription factors in different combinations to give rise to a wide spectrum of expression patterns. Enhanceosome: a higher-order nucleoprotein complex that is formed by the binding of a specific combination of transcription factors to the transcriptional regulatory sequences of a particular gene. General transcription factors: components of the Pol II transcription-initiation complex that are thought to be common to all Pol II promoters. Holoenzyme: a large protein complex that is preformed off of the DNA and contains Pol II and many of the other components of the Pol II transcription-initiation complex. Pol II transcription-initiation complex: a large protein machine that contains dozens of polypeptides and assembles near the transcription start site. Transcriptional activators: class of regulatory proteins that are gene specific and function to increase transcription from target promoters. Transcriptional synergy: when specific combinations of transcription factors give rise to significantly higher levels of transcription than the sum of the additive effects obtained when each factor is assayed individually. During their development and differentiation, plants need to integrate a wide range of tissue, developmental, and environmental signals to regulate complex patterns of gene expression. The regulation of seed-storage protein gene expression, resulting in expression during specific stages of seed development but not in other parts of the plant, is a striking example of tissue and developmental control and is of considerable agricultural importance. Plants also have unique needs and strategies for responding to changes in their environment. When light strikes an etiolated leaf, numerous genes encoding chloroplastic, mitochondrial, peroxisomal, and cytosolic proteins are activated. Similarly, a number of biotic and abiotic stresses cause a battery of genes to be activated as part of the plant-defense/stress response. A major level at which gene expression is regulated is the initiation of transcription, and this is reflected in the percentage of the genome dedicated to transcription factors in plants and other eukaryotes. For example, an analysis of 1.9 Mb of Arabidopsis genomic sequence from chromosome 4 revealed that about 15% of the genes with predicted or known functions were involved in transcription, a percentage similar to what has been found in other eukaryotes (Bevan et al., 1998). In eukaryotic cells, genomic DNA is complexed with proteins to form chromatin. One of chromatin's major roles is to facilitate the packaging of DNA in the nucleus, but the structure of chromatin also leads to a general suppression of gene activity. For gene activation and transcription to occur, the chromatin in the vicinity of the gene must be remodeled to allow access for transcription factors and the recruitment of the RNA polymerase II (Pol II) transcription-initiation complex. Transcription factors play important and diverse roles in gene expression, including chromatin remodeling and recruitment/stabilization of the Pol II transcription-initiation complex. Transcription factors, which come in many shapes and sizes, can be divided into a number of functional classes, with some proteins belonging to more than one class. A major class of transcription factors is activators and repressors. These proteins bind to specific DNA sequences found only in certain promoters and are instrumental in giving rise to gene-specific regulation. A second class of transcription factors are coactivators or corepressors. These proteins mediate the transcriptional effects of specific activators/repressors, in some cases by remodeling chromatin. Whereas this group of transcription factors are typically not able to bind to DNA on their own, they can still be promoter-specific as a result of protein-protein interactions with specific activators and repressors. A third class comprises the general transcription factors, which are important components of the Pol II transcription-initiation complex. A fourth class is architectural transcription factors that are also involved in remodeling DNA, e.g. by inducing bends that facilitate the binding of other proteins to the promoter. In this Update I will address how transcription factors regulate gene transcription in plants, as well as relying on advances in other systems. The focus will be on activators and the importance of combinatorial control. First, I will comment on chromatin, chromatin remodeling, and the Pol II transcription-initiation complex, since it is the recruitment and/or activity of the transcription-initiation complex that is regulated by the gene-specific transcription factors, and this regulation occurs in the context of chromatin. Chromatin has several levels of structural organization, with the basic unit being the nucleosome core, which consists of 146 bp of DNA wrapped around a histone octamer. The presence of nucleosomes affects the accessibility of DNA to other proteins, including transcription factors and the Pol II transcription-initiation complex. Higher orders of chromatin structure are also likely to affect transcription, e.g. by leading to the organization of chromatin into active and silent regions. Exciting progress has been made recently on chromatin remodeling, including the involvement of histone acetylation/deacetylation on nucleosome conformation/stability, and the identification of novel protein complexes that cause nucleosome disruption (for recent reviews, see Cairns, 1998; Struhl, 1998). Histone acetylation was first reported in 1964 and at the time was proposed to play a role in the regulation of transcription (Allfrey et al., 1964). Studies during the next three decades supported this proposal, although until recently the molecular mechanisms involved were not known. Acetylation of histones occurs on Lys residues in the amino-terminal tails that protrude from the surface of the nucleosome. Acetylation neutralizes the positive charge of the histone tails and consequently causes a reduction in their affinity for DNA. This leads to changes in nucleosome conformation and may lead to unfolding of the nucleosome. Thus, acetylation of histones is normally correlated with transcriptional activity by facilitating the access of transcription factors to the DNA, whereas deacetylation of histones is correlated with transcriptional repression. A growing number of histone acetylases and histone deacetylases have been identified recently. Significantly, many of these proteins had already been associated with transcription, with some having been identified previously as components of the Pol II transcription-initiation complex, and others initially identified as coactivators or corepressors. For example, the yeast Gcn5 and mammalian p300/CBP proteins were initially identified as coactivators and were subsequently found to be histone acetylases (Bannister and Kouzarides, 1996; Brownell et al., 1996; Ogryzko et al., 1996). In the case of Gcn5, the histone acetylase activity has been shown to be required for coactivator function in vivo (Candau et al., 1997), and overexpression of Gcn5 leads to increased histone acetylation at promoter regions of genes regulated by Gcn5 (Kuo et al., 1998). Histone acetylation has also been observed in plants (Belyaev et al., 1997) and a maize histone deacetylase has been identified (Lusser et al., 1997). Other forms of chromatin remodeling involving multiprotein complexes with ATP-dependent chromatin remodeling activities have also been observed. A good example is the SWI2/SNF2 complex, which was initially discovered through genetic studies as a transcriptional regulator of specific genes in yeast (Stern et al., 1984; Neigeborn and Carlson, 1984). A number of lines of evidence link the yeast SWI2/SNF2 complex, which has a size of about 2,000,000 D, to chromatin remodeling (for review, see Cairns, 1998). Chromatin remodeling does not occur at promoters normally regulated by the SWI2/SNF2 complex in strains with a defective SWI2/SNF2 complex. In addition, the purified SWI2/SNF2 complex was able to cause chromatin remodeling to occur in vitro. SWI2/SNF2-related complexes have also been identified inDrosophila melanogaster and humans. In D. melangoasterthree additional chromatin-remodeling complexes have been identified, including NURF, which is able to stimulate the binding of a number of transcription factors to chromatin templates (Tsukiyama and Wu, 1996). The identification of distinct multiprotein complexes involved in chromatin remodeling raises important questions regarding the role each plays in transcriptional regulation, how they are targeted to specific promoters, and whether they interact with other types of chromatin-remodeling activities, such as histone acetylases. A simplified schematic representation of the transcription-initiation complex assembled at a typical Pol II core promoter containing a TATA box. Pol II, the general transcription factors IIA, IIB, IID (TBP and TAFs) IIE, IIF, and IIH, as well as a few of the accessory factors (CBP and SRB) are shown. The precise positions of a number of these factors in the transcription-initiation complex remain to be established. Transcriptional activators (ACT) typically bind at specific sites upstream of the core promoter. Interactions between activation domains present on the activators and components of the general transcriptional machinery help to recruit and/or stabilize the transcription-initiation complex. The general transcription factors and some of the accessory proteins were first identified biochemically using in vitro transcription systems. Based on the in vitro studies a stepwise model was proposed for the assembly of the transcription-initiation complex starting with the binding of TFIID to the TATA box (for review, see Roeder, 1996). In this model a major function of transcriptional activators was to facilitate the stepwise assembly of the transcription-initiation complex. However, some of the Pol II in cells has been shown to exist in a large protein complex(es) called the holoenzyme (for review, seeGreenblatt, 1997), suggesting that to a large extent the transcription-initiation complex may already be formed in the absence of DNA. A major role of transcriptional activators may be to recruit the holoenzyme to specific promoters and/or to stabilize the transcription-initiation complex once bound at the core promoter. Until recently, the composition of the transcription-initiation complex was not considered to be a major area of regulation. However, this view is changing with reports of multiple forms of the holoenzyme and the discovery of accessory proteins such as the TAFs. TAFs, which were initially identified as coactivators required by certain activators to function in vitro, also function in core promoter recognition in vivo (Shen and Green, 1997), and TAF250 has histone acetylase activity (Mizzen et al., 1996). Cell-type-specific TAFs have also been identified, e.g. the B-cell- type-specific TAF105 (Verrijzer and Tjian, 1996). The potential for TFIID to serve as a major point of regulation has also been suggested by the discovery of multiple TBPs in some species, including Arabidopsis. In D. melanogaster, in addition to a ubiquitous TBP, a second tissue-specific TBP, TRF (TBP-related factor), has been identified that may be important for neural-specific patterns of gene regulation (Hansen et al., 1997). Recently, a TFIID complex that does not contain TBP has been isolated from human cells (Wieczorek et al., 1998). Surprisingly, this complex, called TFTC (TBP-free TAF-containingcomplex), was shown to be able to substitute for normal TFIID and support transcription from both TATA-box-containing promoters and TATA-less promoters, using an in vitro transcription system. Although these exciting results demonstrate that multiple transcription-initiation complexes exist and offer additional opportunities for the regulation of transcription, it remains likely that the major level of transcriptional control is mediated by transcriptional activators and repressors. Whereas there has been relatively little work on the general transcriptional machinery in plants in comparison with animals and yeast, an increasing number of transcriptional regulatory proteins have been identified in plants. Although transcriptional regulators can be activators or repressors and, in some cases, the same protein can serve both functions, the focus of this Update is on activators. Typically, activators have a modular structure consisting of discrete domains responsible for specific DNA binding, transcriptional activation, and in some cases dimerization and/or other forms of protein-protein interactions. Whereas domain-swap experiments have shown that DNA-binding domains and activation domains can operate independently when fused to a heterologous protein, there is growing evidence that in their native protein context they may sometimes functionally communicate (for review, see Lefstin and Yamamoto, 1998). Structure of the of the yeast protein bound to DNA. 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The that a major function of activation domains is recruitment of the transcriptional machinery to a promoter has support from studies of yeast involving experiments (for review, see and 1997). In these experiments of a DNA-binding to different components of the holoenzyme was to transcription on genes containing the DNA-binding and activators were However, recruitment of the transcriptional machinery may not be the only that activation domains For example, some activation domains may function by the conformation of the transcription-initiation complex assembled at the core facilitating promoter at some during transcription both the and of activation domains to a specific promoter is a major that transcriptional control is which is is that a major eukaryotic transcriptional regulation is combinatorial control. genes are regulated by multiple transcriptional activators by of having a specific of sites in their promoters. a distinct of transcription factors bind to these different sites to give rise to higher-order nucleoprotein complexes that have been called (for a recent review, see 1998). interactions between proteins that form an as well as interactions with components of the general transcriptional can lead to in DNA binding and transcriptional The composition of the assembled at a promoter may in to developmental, or other is to form a number of distinct by using a discrete number of transcription factors in different A to this is through combinatorial a transcription factor can play multiple roles and help regulate different genes expression is by distinct is a growing of evidence from using both in vivo and in vitro for the and importance of in transcriptional regulation. In plants there is also evidence for combinatorial which is The regulation of in maize is one of the in plants for the importance of combinatorial interactions in gene regulation (for recent reviews, et al., 1998). 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The a number of in plants, including to mediate the to a number of environmental as well as the of specific expression patterns during seed gene expression is an important part of (for recent reviews, see and and 1998). A analysis of the sequences required for gene expression has been on a number of such as the gene and and The first to be identified was an sequence in the promoter et al., have been found in the promoters of many including the and are genes that not contain and other sequences have been shown to function in gene expression in some of these promoters. The is similar to a of sequences called the which also contain an core and are present in a number of gene promoters that to different environmental such as and et al., A protein called that to the has been identified et al., may be part of a complex, which and proteins et al., 1998). 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The analysis of transcriptional control in plants will to be an exciting of The progress being made on the of important regulatory proteins, the development of in vitro transcription and the use of genetic for additional using gene will facilitate studies of transcriptional control in plants, which into the mechanisms of and development and lead to agricultural The potential for is by studies of the molecular of to an important since have major on agricultural the identification of genes and the analysis of their expression, an Arabidopsis transcriptional called has been identified et al., 1997), overexpression of which increased the of Arabidopsis plants et al., 1998). similar mechanisms are for in important plants, the use of and/or may help other plants more I to and for their the and and for to use I to the many work or I was to of complex