Promoter eukaryotic structure

Figure 37-9. The eukaryotic basal transcription complex. Formation of the basal transcription complex begins when TFIID binds to the TATA box. It directs the assembly of several other components by protein-DNA and protein-protein interactions. The entire complex spans DNA from position -30 to +30 relative to the initiation site (+1, marked by bent arrow). The atomic level, x-ray-derived structures of RNA polymerase II alone and ofTBP bound to TATA promoter DNA in the presence of either TFIIB or TFIIA have all been solved at 3 A resolution. The structure of TFIID complexes have been determined by electron microscopy at 30 A resolution. Thus, the molecular structures of the transcription machinery are beginning to be elucidated. Much of this structural information is consistent with the models presented here.

The structure and sequence of the catalytic moiety have been determined (06, V6, W6). The enzyme consists of 362 amino acids and 40,638 daltons of the protein predicted by the cDNA sequence. The ADA gene spans 32 kb and consists of 12 exons. The apparent promoter region of the gene lacks the TATA and CAAT sequences often found in eukaryotic promoters and is extremely G/C rich. The location of the ADA gene is on chromosome 20ql2-ql3.11 (Jl). 

Minsky et recently suggested that the emergence of eukaryotes led to a highly crowded environment that may have promoted DNA self-assembly, leading to extremely condensed and thermodynamically stable DNA aggregates such as nucleosomes and solenoid structures of chromatin. 

Qin Y, Hurley LH (2008) Structures, folding patterns, and functions of intramolecular DNA G-quadruplexes foimd in eukaryotic promoter regions. Biochimie 90 1149-1171. Epub 29 Feb 2008. 

Genes can similarly be cloned and expressed in eukaryotic cells, with various species of yeast as the usual hosts. A eukaryotic host can sometimes promote post-translational modifications (changes in protein structure made after synthesis on the ribosomes) that might be required for the function of a cloned eukaryotic protein. 

As already noted, eukaryotic RNA polymerases have little or no intrinsic affinity for their promoters initiation of transcription is almost always dependent on the action of multiple activator proteins. One important reason for the apparent predominance of positive regulation seems obvious the storage of DNA within chromatin effectively renders most promoters inaccessible, so genes are normally silent in the absence of other regulation. The structure of chromatin affects access to some promoters more than others, but repressors that... 

A major goal in recombinant DNA technology is the production of useful foreign proteins by bacteria, yeast, or other cultured cells. Protein synthesis depends upon both transcription and translation of the cloned genes and may also involve secretion of proteins from the host cells. The first step, transcription, is controlled to a major extent by the structures of promoters and other control elements in the DNA (Chapter 28). Since eukaryotic promoters often function poorly in bacteria, it is customary to put the cloned gene under the control of a strong bacterial or viral X promoter. The latter include the X promoter PL (Fig. 28-8) and the lac (Fig. 28-2) and trp promoters of E. coli. These are all available in cloning vehicles. 

Archaebacterial RNA polymerases are very different from their eubacterial counterparts and more closely resemble eukaryotic enzymes both in their subunit complexity and in their amino acid sequences (for review, see Puehler et al., 1989). This view is also reflected in the diversity of the DNA sequences that are used by the transcription apparatus as signals for initiation of transcription, namely, the promoters. Many attempts were made to identify a consensus promoter structure (Zillig et al., 1988). However, as more genes are isolated and characterized, the picture becomes less coherent. Earlier identification of two upstream sequences, box A and box B, located around positions — 30 and + 1, respectively, gave way to two elements —DPE (distal promoter element) and PPE (proximal promoter element)—located - 38 to - 25 and — 11 to — 2, respectively (Reiter et al., 1990). The DPE encompasses the box A sequence TTTA(A or T)A, but the PPE sequence seems to depend more on an (A + T)-rich sequence rather than on a specific DNA sequence. 

In eukaryotic organisms, transcription regulation is a complex process that demands coordinated interaction of several genetic elements. The efficiency of this process mainly depends on the promoter/enhancer sequences, the copy number of the gene, and the structure and elements present at the insertion site in the host s chromatin. On the other hand, the co-transcriptional modifications (capping, splicing, polyadenylation, and transport to cytoplasm) on the primary transcript determine the stability, turnover rate, and translational capacity of the future mRNA. 

Transcriptional control in eukaryotes can be accomplished at several levels. Chromatin structure can control transcription. The formation of so-called hypersensitive sites (sites where the DNA is not bound into nucleosomes) allows protein factors and RNA polymerase to access the DNA. This is necessary for transcription to occur, but hypersensitive sites are not enough. The removal of histone HI allows transcription to occur from a chromatin domain. Some protein factors (for example, TBF) may be bound to a promoter region even if the gene is not being transcribed. TBF also is necessary but not sufficient for transcription. 

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