Module 4 Section 2 Flashcards

1
Q

Processing of RNA Euk vs. Prok

A
  • mRNA subject to constant degradation
  • In prokaryotes, RNA translated before transcription is terminated
  • In Euk, newly transcribed pre-mRNA subject to modifications prior to exit from nucleus
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2
Q

If eukaryotic pre-mRNAs were not post-transcriptionally modified, what would be the consequence?

A
  • enhanced degradation (end caps)
  • Impaired ribosome binding
  • Translation of non-coding DNA sequences
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3
Q

Processing of pre-mRNA

A
  • 5’ capping
  • 3’ polyadenylation
  • splicing
  • nucleotide editing
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4
Q

5’ Capping

A
  • 7-methyl guanosine attached to mRNA through 5’-5’-triphosphate linkage
  • mediated by guanylyltransferase
  • additional methyl groups often added to 2’OH of ribose sugars at the 1st and 2nd nucleotides adjacent to cap, and to N7 on the guanine (the cap one)
  • occurs early in transcription (after addition of 20-30 nucleotides)
  • Mediated by C-term domain (CTD) of RPBI
  • Phosphorylation of CTD recruits specific RNA modifying enzymes (including guanylyltransferase)
  • Cap-binding complex (CBC) recruits mRNA to ribosome
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5
Q

Purpose of 5’ cap

A
  • prevents recognition of the 5’ end by exoribonucleases

- mediates binding of mRNA to the ribosome

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6
Q

3’ polyadenylation steps

A
  1. Pol II transcribes the poly(A) addition site at term end of transcript
    Marked by two sequence elements:
    -highly conserved sequence 5’-AAUAAA-3’ 10-30 nucleotides upstream of cleavage site
    -GT rich region 20-40 nucleotides downstream of cut site
  2. Polyadenylation factors bind poly(A) signal upstream of cleavage site
    -cleavage generates free 3’OH that defines the end of mRNA
  3. “A” residues are added to free 3’OH immediately in Poly(A) polymerase (PAP) catalyzed rxn
    -A-residues are bound by Poly(A) binding proteins (PABP) which protect 3’mRNA from exoribonucleases
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7
Q

mRNA circularization

A
  • 5’ cap and 3’ polyA tail associate with subunits of elF4F to circularize mRNA
    1. mediates ribosome binding
    2. promotes translational efficiency
    3. ensures complete processing
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8
Q

Splicing of Pre-mRNA

A
-exons: typically <1000 nucleotides
Introns:
-50-20000 nucleotides
-do not code for protein
-contain regulatory sequences
-allow for alternative splicing of genes
5'UTR and 3'UTR are NOT spliced out
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9
Q

Splicing

A
  • the controlled act of breaking and joining specific phosphodiester bonds to excise introns
  • cannot reorder RNA (ie. cannot have 2, 1, 3)
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10
Q

Spliceosome

A
  • mediates splicing
  • complex of 5 small nuclear ribonuclear proteins (snRNPs)
  • snRNP = snRNA and protein subunits
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11
Q

Reverse transcription

A
  • generation of cDNA
  • make use of 3’polyA to make sure only mRNA is replicated
  • use polyT sequence as a primer (Oligo dT primer)
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