transcription

Criteria for Molecule-Gene Relationship

  • Molecule-gene characteristics:
    • Replicate: Involves semi-conservative DNA replication, particularly in prokaryotic organisms.
    • Store Information: The genetic material must be capable of retaining the necessary information for cellular function and differentiation.
    • Expression of Information: Involves:
    • Transcription of RNA
    • Translation into polypeptides
    • Folding of polypeptides into functional proteins
    • Increase Variation: Achieved via mutations that can lead to variations in traits.

Goals of the Study

  1. Define cis-element and trans-factor.
  2. Understand the components involved in prokaryotic transcription initiation, both cis-elements and trans-factors.
  3. Describe the sequential steps of prokaryotic transcription: initiation, elongation, and termination.
  4. Learn the distinctions between eukaryotic transcription and prokaryotic transcription.
  5. Recognize the three modifications required to convert eukaryotic pre-mRNAs into mature mRNAs.

Transcription as a Process

  • Transcription is dependent on two types of acting factors:
    • Cis-elements: Specific DNA sequences that are recognized by particular proteins.
    • Trans-factors: Proteins that recognize and bind to specific DNA sequences for transcription.
  • Transcription results in the synthesis of single-stranded RNA.
  • RNA Polymerase: This enzyme synthesizes ribonucleic acids (RNA) using a DNA template.
    • Reads the 3'-5' template strand of DNA to synthesize RNA in the 5'-3' direction.
  • Promoter: A specific DNA sequence located upstream of the transcription starting site essential for transcription initiation.
  • σ Factor (Sigma Factor): A protein crucial for promoter recognition, responsible for recruiting RNA polymerase to the transcriptional start site.

Prokaryotic Transcription Components

  • Components:
    • σ subunit: Part of RNA polymerase
    • Gene: The section of DNA being transcribed
    • Promoter: The starting point for transcription
    • RNA polymerase holoenzyme: The complete enzyme required for synthesizing RNA.

Promoter Cis-Elements in E. coli

  • E. coli promoters contain two consensus sequences recognized by the σ factor:
    • -35 Site: TTGACA, located 35 base pairs before the transcription start site (TSS).
    • -10 Site (Pribnow Box): TATAAT, situated 10 base pairs before the TSS.

Phases of Prokaryotic Transcription

  1. Initiation:
    • This is the rate-limiting step of transcription, heavily regulated by the availability of transcription factors.
  2. Elongation:
    • RNA polymerase synthesizes RNA by forming phosphodiester bonds with nucleotides, creating a RNA strand in the 5'-3' fashion.
    • Example: From the DNA sequence 3'-ATGCAGC-5', the RNA synthesized would be 5'-UACGUCG-3'.
  3. Termination:
    • There are two mechanisms for termination:
      • Hairpin structure: Formed when the transcribed RNA folds back on itself.
      • Rho (ρ) Factor: A protein that facilitates termination by causing the RNA polymerase to detach from the DNA template.

Eukaryotic vs. Prokaryotic Transcription

  • Transcription in eukaryotes differs from prokaryotes in several ways:
    1. Multiple RNA Polymerases: Eukaryotes have three different RNA polymerases, whereas prokaryotes have one.
    2. Reliance on Transcription Factors: Eukaryotic RNA polymerases depend more on transcription factors to locate and bind to the DNA cis-elements during transcription initiation.
    3. Post-Transcriptional Modifications: Eukaryotic mRNA undergoes necessary modifications after transcription to become mature and ready for translation.

Eukaryotic RNA Polymerases

  • Eukaryotes have three distinct forms of RNA polymerase, which transcribe different types of RNA:
    1. RNA Polymerase I (RNAP I):
    • Product: rRNA
    • Location: Nucleolus
    1. RNA Polymerase II (RNAP II):
    • Product: mRNA, snRNA, miRNA, and IncRNA
    • Location: Nucleoplasm
    1. RNA Polymerase III (RNAP III):
    • Product: 5S rRNA, tRNA
    • Location: Nucleoplasm

Elements Required for Eukaryotic Transcription

  • Three main elements are necessary for RNA polymerase to recognize the transcription start site:
    • Promoter: Contains consensus sequences crucial for initiating transcription.
    • Enhancer: A cis-element that assists in gathering transcription factors at the promoter.
    • Enhanceosome: A complex of proteins (activators and coactivators) that help recruit RNA polymerase and other transcription factors to the promoter region, such as TAFs, TBP, etc.

Post-Transcriptional Modifications in Eukaryotic mRNA

  • Special modifications that pre-mRNAs undergo to become mature mRNAs include:
    1. Addition of a 5' cap (7-methylguanylate cap), essential for mRNA stability and translation.
    2. Addition of a 3' Poly-A tail: This involves cleaving the 3' end of the pre-mRNA and attaching a long chain of adenine nucleotides, which protects mRNA from degradation.
    3. Splicing: The process of removing non-coding regions (introns) and joining together coding regions (exons) in the pre-mRNA.

The Splicing Process

  • Exons: Codon sequences that are retained and encode the protein.
  • Introns: Sequences present in the initial RNA transcript that are not expressed in the final protein.
  • Mechanism of Splicing:
    • The removal of introns and the ligation of exons involve ribonucleoproteins (snRNPs), which form a lariat structure during the splicing process leading to the excision of introns and the joining of exons.

Summary of Transcription Processes

  • Synthesis of RNA occurs in a 5'-3' direction using a 3'-5' template strand of DNA, facilitated by RNA polymerase.
  • Transcription progress from:
    • Initiation (Promoter recognition)
    • Elongation (RNA synthesis)
    • Termination (Completion and release of RNA polymerase).