Tarnscription Rna Polymerase

By Alberts

TBP

TATA

By Alberts

By Alberts

Mechanisms of transcription by Pol II 

Structure of RNA polymerase II. Cutaway view, to reveal contents of active center cleft. Surface representation of atomic model, with features colored as follows: clamp, orange; wall, blue; bridge helix, green; active center Mg ion, pink; and remainder of polymerase, gray. A) Transcribing complex, with coding strand of DNA in active center region in turquoise, and RNA in red (PDB 1I6H). B) RNA polymerase II – TFIIB complex, with backbone model of TFIIB in yellow (PDB 1R5U). By Kornberg, (2005) FEBS Letters, 579, 899-903

Structure of RNAPII and interaction of the enzyme with promoter DNA. This schematic representation of the polymerase (shown in orange) emphasizes the way in which the clamp and wall domains restrict access to the active site. Subunits Rpb4 and Rpb7 form a complex (shown in blue) that can dissociate from the core enzyme, and might play a role in helping to determine the position of the clamp domain. The Rpb4–Rpb7 complex may also be involved in interaction with newly synthesized RNA. The narrow configuration of the active site cleft probably requires melting of the transcription start region for the template strand to reach the RNAPII active site (indicated by the red dot). Asturias FJ. 2004. Curr Opin Struct Biol. 14(2):121-9. Review.

RNA polymerase II transcription initiation complex. X-ray and electron microscope structures (upper left) were assembled in a complete transcription initiation complex (lower right). By Kornberg, (2005) FEBS Letters, 579, 899-903

Action of Mediator

Tail

The yeast RNA Polymerase II holoenzyme revealed by electron microscopy and image processing. (A) The extended Mediator contains three distinguishable regions; head (h), middle (m), and tail (t). The globular density embraced by Mediator is identified as RNA polymerase II. The outline of a projection of the previously determined polymerase three-dimensional structure is superimposed (dark line), with the point of attachment of the C-terminal domain (dark circle) and the location of the DNA-binding channel (c) indicated. (B) Tentative subunit organization for the holoenzyme. The model is based on available structural Information and reported physical interactions. The surface of each subunit has been calculated by assuming a globular shape and drawn in scale. Subunits in red have reported homologs in Saccharomyces pombe and, with the exception of Rox3 and Srb6, also in mammalian Mediator. The yellow subunits are specific for Saccharomyces cerevisiae. By Bjorklund & Gustafsson, (2004) Advance in Protein Chem., 67, 43-65

Mediator and its interaction with the basal transcription machinery. The structure of the RNAPII–Mediator complex has revealed the way in which RNAPII interacts with the Mediator complex. As shown, upstream promoter DNA, IIB and TBP are all expected to be located at the interface between polymerase and Mediator. This implies that RNAPII and Mediator cannot arrive at a promoter as a pre-formed complex, but must be recruited independently. Asturias FJ. 2004. Curr Opin Struct Biol. 14(2):121-9. Review.

1. Poised or committed state of the HNF-4 gene

2. Recruitment of CBP, P/CAF, and Brg-1 to the enhancer region and assembly of the RNA pol-II holoenzyme at the proximal promoter region.

3. Unidirectional movement of the DNA-protein complex formed on the HNF-4 enhancer along the intervening sequences and spreading of histone hyperacetylation.

4. Formation of a stable enhancer-promoter complex, hyperacetylation of nucleosomes located at the promoter, remodeling of the nucleosome located at the transcription start site, and release of RNA pol-II from the promoter.

Model Depicting the Sequential Steps Involved in the Formation of an Active Pre-initiation Complex on the HNF-4α Regulatory Region.

Hatzis P, Talianidis I. 2002. Mol Cell 10(6):1467-77.

What is transcription? Transcription is a process that cell selectively copies a particular  portion of its genetic information from DNA ­ a gene ­ to RNA. Gene A TRANSCRIPTION

Gene B TRANSCRIPTION

Transcription  is a process of DNA template­directed RNA  polymerization, or an ordered linkage of ATP, CTP, UTP and GTP  through phosphodiester bonds in 5’ to 3’ direction, with the nucleotide  order determined by complementary base pairing.

Features: It uses one DNA strand as a template. It does not need primers. It synthesizes RNA from 5’ to 3’(unidirectional). DNA duplex unwinds upstream and rewinds downstream of the transcription “bubble”. It involves the function of multiple enzymes and proteins.

Three principal types of RNA polymerases in human  cells synthesize RNAs with different functions Polymerase   Product  I           rRNA

       Regulation         simple

Stability    high

Site of synthesis       nucleolus     nucleoplasm

II

     precursor of mRNA       very diverse

    low

III

     tRNA, small RNA         simple

    high            nucleoplasm

Types of RNA mRNAs rRNAs tRNAs snRNAs

Function code for proteins form the structure of ribosomes and catalyze protein synthesis functions as adaptors between mRNAs and amino acids functions in processes such as pre­mRNA splicing

Lectures are focused on RNA polymerase II (Pol II)­mediated  transcription.

The three stages of transcription: Initiation, elongation and termination. Signals that control RNA polymerases where to start  and stop are encoded in DNA. Gene A

TRANSCRIPTION

Gene B

TRANSCRIPTION

Initiation is the main point at which cells regulate transcription.

Initiation

The selection of a DNA segment to be transcribed is made by the  formation of an initiation complex at the promoter. Promoter: a short region of DNA sequences that specifies the  association of RNA polymerase complex to initiate transcription. Consensus sequences of E. coli promoter

5’………TATTGACA………………TATAAT……Start site         ­35          ­10           +1 The sigma factor in E. coli interacts with the –35 and –10  sequences at the promoters to select genes for the RNA  polymerase to transcribe.