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13 Cards in this Set

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Differentiate between the structures of prokaryotic and eukaryotic mRNAs
Prokaryotic mRNA
Can be polycistronic - it can code for several polypeptides
Is not post-transcriptionally modified
Eukaryotic mRNA
5' cap
3' poly-A tail
Introns are removed
Describe the Post-Transcriptional modifications of eukaryotes mRNA and the significance of these modifications - 5' capping
The cap is composed of a methylated GTP that is linked to the rest of the mRNA by a reverse 5' to 5' triphosphate. This process happens during transcription.
1) A phosphatase removes one phosphate from the 5' end
2) Guanylyl transferase then adds a GMP in a reverse 5' to 5' linkage
3) Guanine - 7 - methyl transferase then adds a methyl group to the 2' - O position on the next to last base on the 5' end. This additonal modification is only found in some mRNAs
Relevance: required for the fficient binding of the ribosome to the mRNA and the subsequent translation of the mRNA into protein.
Describe the Post-Transcriptional modifications of eukaryotes mRNA and the significance of these modifications - 3' Polyadenylation
A typical eukaryote mRNA may possess 100-200 adenines at the 3' end. Polyadenylation occurs in the nucleus following transcription of the mRNA.
1) Cleavage & polyadenylation specificy factor (CPSF) binds to the polyadenylation signal (AAUAAA) in the 3' UTR of the mRNA.
2) Cleavage stimulating factor F (CstF) then binds to a GU-rich region located in the 3' UTR past the point where the 3' UTR is cleaved
3) Cleavage factors then bind to a CA sequence located after the polyadenylation signal and cleave the mRNA at this location. The small cleaved fragment is degraded and the remaining mRNA is polyadenylated.
4) Finally, Poly-A-polymerase (PAP) adds approximately 200 A nucleotides to the new 3' end of the mRNA produced by the cleavage of the 3' URT. The poly-A tail allows the binding of Poly-A binding protein (PABP) which helps in directing translation by the ribosome.
Relevance:
1) Aids mRNA transport out of the nucleus to the cytoplasm
2) Stabilizes mRNAs in the cytoplasm so that they can serve as messages for translation for a longer time
3) Increases the efficiency of the initial steps of translation
Describe the post-transcriptional modifications of eukaryote mRNA and the significance of these modifications - splicing
Both Introns and Exons are expressed (transcribed) in mRNA. The introns are then removed by splicing on both the 5' and 3' ends.
The 5' end (splice donor site) always possesses a GU sequence and the 3' end (splice acceptor site) always has an AG sequence. The conserved branch point sequence, 18-38 bps upstream of the 3' end, always possesses an A nucleotide.
The spliceosome chops the 5' exon-intron junction and forms a lariat structure. The 5' end G folds back and forms a 2' to 5' bond with the A in the branch point sequence. The 3' intron-exon junction is then chopped and the two ends are ligased.
WHY: Allows mRNA to code for multiple proteins or for one protein to be modified into another in a different location.
Spliceosomes
Made up of a number of small molecules called small nuclear ribonucleoproteins (snRNPs
Composed of short-nuclear RNAs (snRNA) & proteins
Bind to the splice junction sites & branch point sequences and facilitate the cleavage, folding and ligation of the pre-mRNA to form mature mRNA.
Assembly requires energy and some of the proteins are RNA helicases, also requiring ATP.
The active site is only created after it's assembly on the mRNA to prevent random splicing. The catalytic site is formed from RNA molecules, not protein.
Alternative Splicing
60% of human genes are spliced in alternate ways, allowing a number of different proteins to be produced from the same gene, such as:
Tissue specific isoforms
Membrane bound & soluble isoforms<br />Alternative intracellular location
Altered function
Alternative splicing can occur due to intron sequence ambiguity (chance) or directed by proteins that bind to splice sequences and exert either positive or negative control.
Systemic Lupus Erythematosis
An autoimmune disease
Symptoms: extreme fatigue, arthritis, fever, skin rashes, kidney problems
Mechanism: anti-bodies cross react with the U1 RNA component of the spliceosome, preventing normal mRNA splicing
Treatment: Symptoms only
Beta thalassemia
Reduced synthesis of beta hemoglobin chain that results in microcytic hypochromic anaemia
Mechanism: Generate additional splice sites with the mRNA introducing frame shifts of premature stop codons and the production of abnormal beta-globin proteins
Limb girdle muscular dystrophy (LGMD)
Symptoms: Weakness and wasting restricted to limb musculature.
Mechanism: a mutation in the calpain 3 gene generates a new splice site within exon 16. Results in a shorter calpain 3 mRNA and a defective protein.
A - I (inosine) editing by Adenosine Deaminases acting on RNA enzyme (ADAR)
Mechanism: Complementary RNA sequences occur in an exon & intron forming a ds RNA structure, attacting the ADAR enzyme.
ADAR1 - liver, required for normal erythrocyte development
ADAR2 - required for normal brain development involving learning and memory
C - U editing in Apolipoprotein-B mRNA by the action of Cytidine Deaminase
In the bowel, a C to U change creates a stop codon. This keeps the protein, made in the liver, from acting until it's in the higher pH small intestine.
Trypanosomes &amp; Leishmania
Parasites who use editing to change form, allowing them to hide first in neutrophils and later in macrophages.