--- TRANSCRIPTION FACTORS AND RNA POLYMERASE IN EUKARYOTES AND PROKARYOTES 1. RNA POLYMERASE IN PROKARYOTES VS. EUKARYOTES A. Prokaryotic RNA Polymerase - In pr
--- TRANSCRIPTION FACTORS AND RNA POLYMERASE IN EUKARYOTES AND PROKARYOTES 1. RNA POLYMERASE IN PROKARYOTES VS. EUKARYOTES A. Prokaryotic RNA Polymerase - In prokaryotic organisms (such as bacteria), transcription is carried out by a single RNA polymerase enzyme . - This RNA polymerase does not require additional transcription factors to initiate transcription. - Instead, the enzyme recognizes specific promoter sequences with the help of a sigma (σ) factor that enables the enzyme to locate the transcription start site. - Once transcription begins, the σ factor is released , allowing the core enzyme to elongate the RNA transcript. B. Eukaryotic RNA Polymerase - Eukaryotic transcription is more complex and requires three distinct RNA polymerases : RNA Polymerase I – Synthesizes rRNA (except 5S rRNA). - RNA Polymerase II – Synthesizes mRNA, some small nuclear RNAs (snRNAs), and microRNAs. - RNA Polymerase III – Synthesizes tRNA, 5S rRNA, and other small RNAs. - RNA Polymerase II, which transcribes mRNA , requires multiple transcription factors to recognize and bind to promoter regions, initiate transcription, and regulate gene expression. 2. TRANSCRIPTION FACTORS IN EUKARYOTES A. Definition and Role - Transcription factors (TFs) are regulatory proteins that assist RNA polymerase in initiating transcription by recognizing and binding to promoter regions. - They help in unwinding DNA, stabilizing RNA polymerase binding , and regulating gene expression . B. Transcription Factors for RNA Polymerase II - RNA Polymerase II requires six essential transcription factors , which are collectively called General Transcription Factors (GTFs) . - These factors are necessary for the initial stages of transcription and help in: Recruiting RNA Polymerase II to the promoter region . - Unwinding DNA at the transcription start site . - Stabilizing the transcription complex . - Regulating the initiation process . C. The Six General Transcription Factors for RNA Polymerase II Each transcription factor has specific roles in the formation of the transcription initiation complex : - TFIID (TATA-Binding Protein, TBP) Structure & Function : Contains TBP (TATA-Binding Protein) , a monomeric protein that recognizes the TATA box , a conserved promoter sequence. - TBP binding to the TATA box is the first step in transcription initiation . - TFIID also contains TAFs (TBP-associated factors) , which help regulate transcription and provide specificity to different promoters. - Key Role : Initiates transcription by binding to the TATA box in the regulatory region of the gene. - TFIIA Structure & Function :A multimeric protein composed of three subunits. - It stabilizes the binding of TFIID (TBP) to the TATA box. - Enhances RNA polymerase II recruitment to the promoter. - Key Role : Ensures stability of the transcription initiation complex. - TFIIB Structure & Function :A monomeric protein that binds to TFIID (TBP) . - Helps position RNA Polymerase II correctly at the transcription start site. - Recruits TFIIF and RNA Polymerase II to form the pre-initiation complex. - Key Role : Bridges the interaction between TBP and RNA Polymerase II , ensuring accurate transcription start site selection. - TFIIF Structure & Function :A multimeric protein with two subunits. - Binds tightly to RNA Polymerase II , preventing it from binding non-specifically to DNA . - Helps RNA Polymerase II bind specifically to the correct promoter . - Key Role : Prevents random DNA binding and stabilizes RNA Polymerase II during initiation. - TFIIE Structure & Function :A multimeric protein with two subunits. - Recruits TFIIH , an essential transcription factor with enzymatic activity. - Plays a role in DNA unwinding , allowing the transcription machinery to access the DNA template. - Key Role : Facilitates DNA opening by recruiting TFIIH . - TFIIH Structure & Function :A multimeric protein with 12 subunits. - Contains helicase activity , which unwinds DNA at the transcription start site . - Phosphorylates RNA Polymerase II , activating it for elongation. - Involved in nucleotide excision repair (NER) , a DNA repair mechanism that corrects transcription errors. - Key Role : Unwinds DNA to allow transcription initiation. - Phosphorylates RNA Polymerase II to trigger the elongation phase . - Plays a role in DNA repair . 3. FORMATION OF THE TRANSCRIPTION INITIATION COMPLEX The process of transcription initiation follows these steps: - TFIID (TBP) binds to the TATA box , marking the transcription start site. - TFIIA stabilizes TFIID binding . - TFIIB bridges TBP and RNA Polymerase II , recruiting the enzyme to the promoter. - TFIIF binds to RNA Polymerase II , preventing it from interacting with non-specific DNA sequences. - TFIIE recruits TFIIH , which begins DNA unwinding . - TFIIH unwinds DNA and phosphorylates RNA Polymerase II , allowing it to transition from initiation to elongation. Once this complex is assembled and RNA Polymerase II is activated, transcription begins. 4. DNA REPAIR DURING TRANSCRIPTION - TFIIH is involved in transcription-coupled DNA repair . - It recruits the nucleotide excision repair (NER) machinery , which detects and corrects errors in the DNA template strand . - This process ensures accurate RNA synthesis and prevents mutations. 5. SUMMARY TABLE OF TRANSCRIPTION FACTORS FOR RNA POLYMERASE II 6. CONCLUSION - Prokaryotic transcription is simple , requiring only a single RNA polymerase and a sigma factor . - Eukaryotic transcription is complex , requiring multiple transcription factors to properly regulate gene expression. - RNA Polymerase II requires six transcription factors (TFIID, TFIIA, TFIIB, TFIIF, TFIIE, and TFIIH) to initiate transcription. - TFIIH is unique , as it has helicase activity for DNA unwinding and is involved in DNA repair . - The correct assembly of these factors ensures precise and efficient transcription in eukaryotic cells. --- GENE STRUCTURE AND TRANSCRIPTION IN PROKARYOTES & EUKARYOTES 1. GENE STRUCTURE Genes in both prokaryotic and eukaryotic organisms contain several important regions that regulate transcription and determine which proteins will be synthesized. A. Structure of a Prokaryotic Gene - Prokaryotic genes are arranged in operons , meaning multiple genes may be transcribed together. - These genes produce polycistronic mRNA , which codes for more than one protein . - Key Regions of a Prokaryotic Gene : Upstream Regulatory Region :Contains regulatory sequences such as promoters and operators . - The promoter is the binding site for RNA polymerase and contains conserved sequences at -10 and -35 positions. - Promoter Region : Located at positions -10 and -35 upstream of the transcription start site. - -10 region (Pribnow box) : TATAAT sequence (helps in melting/opening the DNA). - -35 region : Contains TTGACA sequence (recognized by RNA polymerase). - Structural Gene :Codes for the proteins required by the cell. - Multiple genes are often transcribed together in operons (e.g., the lac operon ). - Downstream Terminator Region :Signals the end of transcription and releases the mRNA. B. Structure of a Eukaryotic Gene - Eukaryotic genes produce monocistronic mRNA , meaning one gene codes for one protein . - Key Regions of a Eukaryotic Gene : Upstream Regulatory Elements :Includes several promoter elements and enhancers that regulate transcription. - Promoter Region :Contains multiple regulatory sequences : TATA Box (Hogness Box) : Located around -25 to -30 , recognized by TBP (TATA-binding protein) . - CAAT Box : Found upstream of the TATA box (~ -80), enhances transcription efficiency. - GC Box : Located at -100 to -150 , plays a role in gene regulation. - Coding Region :Consists of exons (coding sequences) and introns (non-coding sequences) . - Introns are removed during mRNA processing (splicing). - Downstream Terminator Region :Signals RNA polymerase to stop transcription. - Includes a polyadenylation signal (AAUAAA) that leads to mRNA cleavage and polyadenyl