Lecture 6

• Post-transcriptional regulation

  Modifications of the RNA transcript after it is produced by RNA polymerase II (Pol II)
• Post-transcriptional modifications

  • Capping
  • Splicing
  • Polyadenylation
• 5' Capping of mRNA

  1. RNA Triphosphatase removes gamma phosphate

  2. Guanylyltransferase adds GMP

  3. Methyltransferase adds methyl group
• Biological significance of 5' capping

  • Distinguishes Pol II transcripts
  • Stabilizes RNA against exonucleases
  • Necessary for efficient translation and further RNA processing
• RNA Splicing

  1. Removal of non-coding sequences (introns)

  2. Joining of coding sequences (exons)
• Spliceosome

  Complex of proteins and RNA that carries out the splicing process
• 3' End Processing

  1. Cleavage of RNA at specific site

  2. Addition of poly(A) tail
• Functions of Polyadenylation

  • Enhances mRNA stability
  • Facilitates mRNA export from nucleus
  • Plays role in translation initiation
• mRNA Export from Nucleus to Cytoplasm

  1. Interaction of mRNA with nuclear pore complexes

  2. Facilitated by export proteins recognizing cap and poly(A) tail
• mRNA Decapping and Degradation

  1. Removal of 5' cap by decapping enzymes

  2. Degradation via 5' to 3' or 3' to 5' exonucleases
• Regulatory significance of post-transcriptional modifications

  • Protect and prepare mRNA for translation
  • Provide mechanisms to rapidly alter protein synthesis without new transcription
• Introns were first identified in the rRNA of the protozoan Tetrahymena
• Mechanism of Splicing

  1. U1 snRNP binds 5' splice site

  2. U2 snRNP binds branch site

  3. Lariat formation and exon joining
• Alternative Splicing

  Allows single gene to produce multiple protein variants
• Improper splicing can lead to diseases such as Spinal Muscular Atrophy (SMA) and some forms of cancer
• Polyadenylation

  Process of adding a tail of adenine nucleotides to the 3' end of a pre-mRNA transcript
• Key components of polyadenylation machinery

  • CPSF
  • CF
  • CstF
  • PAP
  • PABP
• Process of Polyadenylation

  1. Signal and cleavage site recognition

  2. Poly(A) tail synthesis
• Functions of Poly(A) Tail

  • Facilitates nuclear export
  • Enhances translation efficiency
  • Protects mRNA from degradation
• Defects in polyadenylation machinery can lead to diseases due to improper mRNA processing