Transcription-requiring complex

The formation of a transcriptionally active complex requires the interaction of all transcription cofactors with their respective specific DNA sequences. Once they have bound to their specific sequences, it is on these that the remaining elements of the complex that do not interact directly with the DNA are assembled (Chin 1995 Filardo 2002). The elements of the complex that do not come into direct contact with DNA have their own specificity of interaction with the remaining proteins of the complex. Therefore, they include important restrictions so that a fully active transcriptional complex can be assembled with difficulty on a receptor dimer that has incorrectly recognized a HRE. Indeed, an incorrect interaction can imply a noticeable degree of transcription inhibition. 

In contrast to the procaryotes, where the o -holoenzyme of the RNA polymerase can initiate transcription without the aid of accessory factors, the eucaryotic RNA polymerase requires the help of numerous proteins to begin transcription. These proteins are termed basal or general initiation factors of transcription. Together with RNA polymerase II, they participate in the basal transcription apparatus. The various components must associate in a defined order for the formation of a transcription-competent complex, from which a low level of transcription is possible. An increase in the basal transcriptional level requires the effect of specific transcriptional activators, which bind cognate DNA sequences at a variable distance from the promoter. The transcriptional activators themselves require the aid of further protein factors, known as coactivators (see 1.4.3.2), in order to attain full stimulatory activity. 

Coactivator Protein Complexes Most transcription requires the presence of additional protein complexes. Some major regulatory protein complexes that interact with Pol II have been defined both genetically and biochemically. These coactivator complexes act as intermediaries between the DNA-binding transactivators and the Pol II complex. 

Eukaryotic cells contain multiple copies of the 5S rRNA gene. Unlike other eukaryotic rRNA genes, the 5S rRNA genes are transcribed by RNA Pol III. Two control elements, an A box and a C box, lie downstream of the transcriptional start site. The C box binds TFIIIA which then recruits TFIIIC. TFIIIB now binds and interacts with RNA Pol III to form the transcription initiation complex. Transcription produces a mature 5S rRNA that requires no processing. 

The rRNA promoter consists of a core element which straddles the transcriptional start site (designated as position +1) from residues -31 to +6 plus an upstream control element (UCE) about 50-80 bp in size and located about 100 bp upstream from the start site (i.e. at position -100 Fig. 4b). A transcription factor called upstream binding factor (UBF) binds both to the UCE as well as to a region next to and overlapping with the core element. Interestingly, TATA box binding protein (TBP see Topic G6), also binds to the rRNA promoter (in fact, TBP is required for initiation by all three eukaryotic RNA polymerases). The UBF and TBP transcription factors interact with each other and with RNA Pol I to form a transcription initiation complex. The RNA Pol I then transcribes... 

Fig. 10.8 Above, import of the transcription factor NF-AT4 into the nucleus. In activated cells, import is initiated by calcineurin-mediated dephosphorylation of NF-AT4. Dephosphorylation unmasks the nuclear-localization signal (NLS), and at the same time blocks the nuclear export signal (NES). The NES is recognized by the exportin protein (Crml). Nuclear export is an active process. Moreover, nuclear export requires rephosphorylation of the NF-AT4 transcription factor. It is indicated that dephosphorylation by calcineurin and nuclear export are mutually exclusive, because calcineurin and Crm 1 compete for a common binding site on NES. When NES binds to Crml, NT-AT4 is exported from the nucleus, and when calcineurin binds to NES, NF-AT4 remains in the nucleus and forms a transcriptionally active complex. Below, how the extent of dephosphorylation controls the transcriptional activity of NF-AT4. When NF-AT4 is fully phosphorylated, NLS is hidden and the transcription factor remains in the cytoplasm. When NF-AT4 is only partially dephosphorylated, NLS is exposed and can interact with importin a/b which promote nuclear import, and at the same time, NES can interact with the exportin Crml, which promotes nuclear export. The consequence is that the transcription factor shuttles between the nucleus and the cytoplasm and is not transcriptionally active. In order to become transcriptionally fully active, NF-AT4 must be completely dephosphorylated. This prevents export from the nucleus by blocking NES, and may increase the affinity of the transcription factor for DNA by exposure of its trans-activating domain (TAD). (The entire scheme is reproduced with permission of Drs Patrick G. Hogan and Anjana Rao and Nature from Fig. 1 in ref. 68.)...

Like replication, transcription requires separation of the duplex DNA strands and uses a polymerase to copy the template DNA strand. For transcription, the polymerase is RNA polymerase II, which binds to sequences in the regulatory region of the gene called the promoter. Promoters occur approximately 100 bases upstream (i.e., at the 5 end) from the initiation site of transcription where the first ribonucleotide unit is paired with the template (uracil pairs with adenine). Promoters are usually rich in thymine and adenine in repeating patterns and have been referred to as a TATA box. Initiation of transcription requires many protein cofactors to bind to RNA polymerase to form the active initiation complex. Other regions of DNA known as enhancers may interact with the initiation complex to stimulate or repress transcription. Regulation of transcription is the primary mechanism cells use to control gene expression. ... 

When all these elements are bound to DNA, the basal transcription apparatus complex is formed and can transcribe DNA slowly. Other factors are required for fast, efficient mRNA synthesis. 

The mode of action of Smad 4 clearly differs from that of the other members of the Smad family. Smad 4 binds to phosphorylated R-Smads and forms trimeric complexes composed of two R-Smad molecules and one Smad 4 molecule. These complexes translocate to the nucleus, where they bind to related DNA elements and activate the transcription of target genes. The mechanism of transcription regulation by Smads is complex and includes both positive and negative influences. Generally, Smad-dependent regulation of transcription requires the interaction with other transcription factors, such as members of the FoxH 1 family of forkhead transcription factors, the Vitamin D receptor and the c-Jun transcription factor, among others (review Attisano et al., 2001). Futhermore, Smads can interact with coactivators and corepressors of transcription and thereby recruit, e. g., histone acetylase activity or histone deacetylase activity to chromatin. 

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