Eukaryotic_transcriptional_regulation

Transcriptional Regulation in Eukaryotes

RNA Polymerases

• Eukaryotes have three main RNA polymerases:

  • RNA Polymerase I (Pol I): Transcribes ribosomal RNA (rRNA).

  • RNA Polymerase II (Pol II): Main enzyme for transcribing messenger RNA (mRNA), many micro RNAs, and some noncoding RNAs.

  • RNA Polymerase III (Pol III): Transcribes transfer RNA (tRNA) and some smaller rRNAs.

Promoters and Core Elements

• All RNA Polymerases utilize TATA Binding Protein (TBP) to bind to gene promoters.

• TATA Box:

  • Located approximately 30 nucleotides upstream of the transcription start site.

  • Consensus sequence: TATAAA.

  • TBP is recruited to all genes, regardless of the presence of the TATA box.

• Other conserved promoter elements include:

  • BRE Element:

    • Located approximately 30 to 37 nucleotides upstream.

    • Recognized by transcription factor IIB (TFIIB).

  • Initiator Sequence:

    • Directly at the transcription start site (from -1 to +4).

  • Downstream Promoter Element (DPE):

    • Located about 28 to 32 nucleotides into the gene sequence.

Transcription Factors

• General Transcription Factors (GTFs) are integral for transcription:

  • TFIIB: Recognizes and binds the BRE element.

  • TFIID: Contains TBP and binds the TATA box.

  • Other GTFs include TFIIA, TFIIE, TFIIF, and TFIIH.

• Mediator Complex:

  • A 20-protein complex that facilitates the interaction between RNA Polymerase II and other transcription factors, especially those bound distantly.

  • Enhancers interact with the mediator to promote transcription.

Enhancers and Distal Control Elements

• Enhancers are sequences that can be located far from the promoter and are crucial for gene expression:

  • They recruit specific transcription factors, which might bind to various locations in the enhancer sequence.

  • Mediator bridges enhancers to the promoter by bending the DNA.

• Combinatorial Control:

  • Gene expression requires multiple transcription factors (activators and repressors) to be present.

  • Activators and repressors interact with the transcription machinery, influencing expression levels.

Examples of Gene Regulation

• Even Skipped Gene (Eve) in Drosophila:

  • Expression is regulated by gradients of transcription factors (e.g., Bicoid and Hunchback as activators; Giant and Krüppel as repressors).

  • Specific enhancers control the formation of the characteristic 7 stripes of Eve expression.

• Insulators:

  • Sequences that block enhancer signals from activating non-target promoters.

  • Example: CTCF protein binds to insulator sequences to separate gene A's promoter from gene B's enhancer activity.

Genetic Imprinting Example

• IGF2 Gene:

  • Involved in imprinting; the paternal allele is expressed while the maternal allele is silenced due to methylation of the insulator sequence.

Studying Gene Regulation

• Deletion Analysis:

  • Employs reporter genes to identify necessary promoter and enhancer sequences.

  • Example: Reporter genes such as luciferase or GFP are used to observe expression patterns when studying promoter function.

• DNA Footprinting:

  • Method used to determine where transcription factors bind on DNA.

  • DNase I exposure results in protected regions that indicate binding sites of proteins on the promoter sequences.