Nuclear receptors
as the steroid hormones, and those that bind as heterodimers to direct repeat sites.
Activation and repression of transcription
The mechanism of action of transcription factors is complex. In general, the basal transcription machinery of a gene is assembled around a region of DNA upstream of the coding sequence, called the core promoter. Its sequence is just sufficient to specify unregulated (basal) transcription. Core
promoters that possess a TATA element bind TBP (TATA-binding protein), which nucleates the binding of the other subunits (termed general transcription factors) to form a pre-initiation complex that includes RNA polymerase II. Activation of this complex and the initiation of transcription by RNA polymerase occur when transcriptional regulators bind at response elements further upor downstream. The interaction of these sites with the pre-initiation complex takes advantage of the flexibility of DNA and involves yet more accessory proteins that mediate interactions between the regulatory and general transcription factors. The regulation of transcription by nuclear receptors follows this pattern, relying on key accessory proteins termed coregulators.
Coactivators
Ligand-bound nuclear receptors attached to DNA at a response element activate transcription by the recruitment of coactivators. These associate with receptors mostly via the amphipathic AF-2 regions of the LBD (see Figure 10.4). Coactivators possess a nuclear receptor box containing a number
of LxxLL motifs, which bind to a hydrophobic surface on the receptor. This surface is exposed only when ligand is bound due to the rearrangement of the AF-2 region (helix 12), illustrated in Figure 10.5 for the LBD of RXR.
The effect of coactivator binding is to increase the rate of transcription.
This requires interactions with components of the transcriptional machinery (the general transcription factors) and, at the same time, the opening up of the chromatin structure to provide access. Rearrangement of chromatin is brought about by ATP-dependent chromatin remodelling complexes and also by acetylation of histones. This leads to the unwinding of DNA from
its compact structure on nucleosomes; (this theme recurs: see Figure 14.3, page 423 and Figure 22.5, page 710). Some coactivators possess histone acetyltransferase (HAT) activity, others form complexes that activate chromatin remodelling and yet others interact indirectly with the general transcription factors. Figure 10.9a shows a simplified activation scheme for a steroid hormone receptor (GR). The ligand-bound receptor recruits coactivators principally through its AF-2 domain (for example SRC-2, CBP/p300, and p/CAF).
A complex is formed that possesses HAT activity and that interacts with
The TATA element is a highly conserved DNA sequence present in the promoter region of most genes. It binds
transcription factors and histones, the binding of the one acting to block the binding of the other. TATA boxes, usually located 25 bp upstream to the transcription
site, are involved in the initiation of transcription by RNA polymerase.
Many genes lack a TATA box, its place taken by an initiator element
or downstream core promoter. Such TATA-less promoters nonetheless always involve the TATAbinding protein (TBP) which binds without sequence specificity.
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Signal Transduction
FIG 10.9 A simplified scheme of transactivation by glucocorticoid and vitamin D receptors.
the general transcription factors. A chromatin remodelling factor (BRG-1) is recruited to the AF-1 domain.
Corepressors
Nuclear receptors that bind as heterodimers to direct repeat half-sites are commonly nucleus-resident and may be located on DNA even in the
absence of ligand (e.g. thyroid hormone receptors, but not steroid hormone receptors). In this state the AF-2 sites attach corepressors, the converse of coactivators, which act to repress the level of basal transcription. For example, the corepressors NCoR and SMRT recruit histone deacetylases that stabilize chromatin, keeping the DNA strands tightly wound on nucleosomes. They also interact with the general transcription factors in an inhibitory fashion (Figure 10.9b). The binding of ligand to the LBD induces the conformational change, in particular the rearrangement of H12 in AF-2 as discussed above, which releases the corepressor and allows it to be replaced by a coactivator, such as SRC-1. This leads to the recruitment of other coactivators with HAT activity, such as pCAF and p300/CBP. Also, TRAP/DRIP coactivators form
a complex that interacts directly with the general transcription factors. Because of the opposing effects of corepressors and coactivators, the effect
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Nuclear receptors
of ligand binding is to increase transcription from a fraction of the basal level (corepressor bound), to many times the basal level (coactivator bound), generating a response with a large dynamic range.
Transrepression
In the absence of ligand, nucleus-resident receptors bind corepressors which lead to the repression of transcription at a particular response element. But some receptors can work in the reverse sense, so that in the absence of ligand, transcription proceeds constitutively, but in its presence it is inhibited. This is transrepression and it is less well understood than transactivation. Here, we take the thyroid hormone receptor (TR) as an example.
Transactivation by TR involves the binding of RXR-TR heterodimers to thyroid hormone response elements and the activation of transcription in the presence of the hormone T3 as described above. In the absence of ligand the receptor represses transcription by recruitment of corepressors. However, there are also negative response elements to which the receptor can bind. Here, the receptor activates transcription in the absence of T3, but represses it when
T3 is bound, silencing target genes that encode thyroid stimulating hormone (TSH) or thyrotropin-releasing hormone (TRH). The mechanism involves recruitment of HDAC activity and there is a requirement for corepressors such as SMRT. Curiously however, in the absence of ligand, the corepressor acts as a coactivator and this suggests that coregulator function may finally depend on the type of response element involved (positive or negative).
Regulatory networks
Because a single receptor, such as TR, can bind different types of response element, different nuclear receptors may vie for the same or overlapping target sequences. Moreover, they may have common ligands. All this can lead to crosstalk between different receptor mechanisms. Other ways in which receptors interfere with each other include the sequestration of coregulators or recruitment of corepressors at nearby sites. Thus, some orphan receptors can be viewed as constitutive repressors or controllers of basal transcription through corepressor recruitment and the consequent HDAC activity and interaction with the general transcription factors. Conversely, other orphans may increase responsiveness of particular nuclear receptors.
Such crosstalk forms the basis of regulatory networks that underpin important physiological functions. These transcriptional control systems are often complex. An example is provided by the nuclear receptors that regulate the breakdown of cholesterol in the liver to form bile acids. The first and rate-limiting enzyme in the catabolism of cholesterol is a member of the cytochrome P450 family CYP7A1 (cholesterol 7 -hydroxylase). A negative
Cytochrome P450 proteins. These haem proteins are oxidoreductases that catalyse the oxidation of many endogenous and exogenous
hydrophobic compounds, such as fatty acids, sterols, prostaglandins, leukotrienes, and a wide range of xenobiotics. Some 40 different P450 protein families have been identified on the basis of their sequences. Humans express 57 different proteins. P450 proteins are classified with the prefix CYP.
Note: the CYPs are not related to the cyclophilins that have been assigned a similar prefix, Cyp. Nor is SHP related to SHP-1 and SHP-2, Src homology domain phosphatases (see page 644).
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