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Gene Regulatory DNAs and Proteins Transcriptional control is the most important type of control for most genes. Both bacteria and eucaryotes use gene regulatory proteins (activators and repressors) to regulate their genes. A gene regulatory protein recognizes a specific DNA sequence because the surface of the protein is extensively complementary to the special surface features of the double helix in that region of binding. The binding involves many hydrogen and ionic bonds and hydrophobic interactions. It is the distinctive features of hydrogen bond donors, acceptors and methyl groups that these proteins recognize and these features are most varied at the major groove which is why binding occurs at the major groove. Most eucaryotic gene regulatory proteins do not act alone but rather act as parts of complexes composed of often many polypeptides. These complexes often assemble only in the presence of the appropriate DNA sequence. Gene regulatory proteins that do not themselves bind DNA but assemble on DNA bound gene regulatory proteins are often called coactivators or corepressors depending on their effect on transcription initiation. Typically the assembly of a group of regulatory proteins on DNA is guided by a few relatively short stretches of nucleotide sequence but in some cases a more complicated protein-DNA structure called an enhancesome is formed. One of the proteins in an enhancesome is called an "architectural protein" that bends the DNA and thereby promotes the assembly of the other enhancesome proteins. Many DNA regulatory proteins have similar structural motifs. One such motif is called the leucine zipper which consists of 2 alpha helices joined to each other by interactions between hydrophobic amino acid side chains that extend from one side of each helix. Just beyond the dimerization interface the 2 alpha helices separate form each other to form a Y shaped structure which allows their side chains to contact the major groove of DNA. The dimer thus grips the double helix like a clothespin. Many gene regulatory proteins like leucine zipper proteins can associate with nonidentical partners to form heterodmimers (rather than homodimers) composed of 2 different subunits which can greatly increase the binding specificities that these proteins display. Another major motif is the helix-turn-helix which involves 2 alpha helices connected by a short extended chain of amino acids (the "turn"). One of the helixes has a C-terminal called the recognition helix which fits into the major groove of DNA. Its amino acid side chains differ from protein to protein and plays a crucial part in recognizing the specific DNA sequence to which the protein binds. The helix-turn-helix proteins bind as symmetric dimers to DNA sequences. Another motif is the helix-loop-helix which consists of a short alpha helix connect by a loop to a second longer alpha helix. This helix-loop-helix structure binds both to DNA and to a second HLH protein. The second HLH protein can be the same (creating a homodimer) of different (creating a heterodimer). Another important DNA binding protein motif is called zinc fingers add one or more zinc atoms as structural components. One type that was dicovered to activate the transcription of a eucaryotic robosomal RNA gene consists of an alpha helix and a B sheet held together by zinc. Zinc fingers are often clustered in 3 and typically recognize GC rich sequences. Eucaryotic gene activator proteins increase the rate of transcription by attracting, positioning and modifying general transcription factors and RNA polymerase II at the promoter. Enhancers The regulation of transcription in eucaroytes differs in a number of ways from that of bacteria. First, proteins can regulate transcription of a gene in eucaryotes by binding to DNA sequences thousands of base pairs away from the promoter which they influence. These DNA sequences to which activators bind are sometimes called enhancers since they enhance the rate of transcription. The model for how such enhancers work is that the DNA between the enhancer and the promoter loops out to allow the activator proteins bound to the enhancer to come into contact with proteins bound to the promoter like RNA polymerase or a general transcription factor. A typical animal gene is likely to contain several enhancers that can be located in 5' and 3' regulatory regions, as well as within introns. Each enhancer is responsible for a subset of the total gene expression pattern; they usually mediate expression within a specific tissue or cell type. A typical enhancer is around 500 bp in lenght and contains on the order of ten binding sites for at least 3 different sequence-specific transcription factors, most often two different activators and one repressor. Locus Control Region (LCR) A LCR is an element that confers tissue-specific high level expression to linked genes, presumably by overriding suppressive effects of flanking DNA sequences. Experimentally, a LCR is an element that gives transgene copy number dependent and integration site-independent expression to linked genes in transgenic mice. The critical difference between LCRs and enhancers is that LCRs enable position-independent expression in transgenic mice, which is reflected in the copy number dependency of the construct, whereas enhancers do not have this property. Insulator DNAs Insulator DNA prevent enhancers associated with one gene from inappropriately regulating neighboring genes. Insulators are typically 300 bp to 2 kb in lenght and often contain clustered binding sites for large zinc finger proteins, such as Su(Hw) and CTCF. They selectively block the long-range interaction of a distal enhancer with a proximal target promoter when positioned between the two. Insulators were first identified at gene boundaries where they prevent cis-regulatory sequences in one gene from inappropriately interacting with neighboring loci. Silencers Silencers such as those of the immunoglobulin k lgith chain gene are located adjacent to their promoter. In constrast, the sileners of the TNF gene are located at a distance from their promtoer. Although silencers have been functionally demonstrated in numerous vertebrate genes, the question of how trnascriptional silencers act against the target promoter is still unanswered. Chromatin remodelling and Modifying Complexes Many gene activator proteins bind and thereby recruit histone acetyl transferases (HATs) also known as histone acetylases as well as ATP dependent chromatin remodeling complexes.which results in greater accessibility to DNA which aides the assemble of general transcription factors and the RNA polymerase at the promoter. Eucaryotic gene repressor proteins can inhibit transcription in a variety of ways such as by 1) competitive binding to the same regulatory DNA sequence as a gene activator, 2) masking the activation surface of an activator, 3) interaction with the general transcription factors such as by blocking their assemble, 4) recruiting repressive chromatin remodeling complexes and 5) recruitement of histone deacetylases. Control of the Regulatory Proteins: Regulatory proteins are themselves regulated by a whole set of other proteins. In addition, regulatory proteins are influenced by signals from outside the cell, which can make them active or inactive in several different ways such as the following:
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