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Gene Expression > Post-transcriptional control of gene expression > Flashcards

Post-transcriptional control of gene expression Flashcards

1
Q

What is added to eukaryotic mRNAs post-transcriptionally ?

A

Cap and poly A tail, they are not encoded in the genome.

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2
Q

What is pre-mRNA?

A

The unfinished messenger RNA or precursor messenger RNA is known as pre-mRNA.

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3
Q

What events are coupled to transcription?

A

*Capping
*Splicing
*Polyadenylation
*Editing

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4
Q

How is the 5’ m7G cap synthesised?

A

1) GpppN structure
2) methylation

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5
Q

What is the structure of the 5’ m7G cap?

A
  • All RNA pol II RNAs
  • RNA initially contains triphosphate at 5’ end
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6
Q

What is the importance of methylation in the synthesis of the 5’ m7G cap?

A

Methylation alters chemical behaviour of base

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7
Q

What are the Functions of the m7G cap?

A
  • Protects mRNA from degradation by 5’-3’ nucleases
  • Facilitates splicing
  • Facilitates export from the nucleus
  • Critical for translation of most mRNAs
  • Functions mediated through protein binding
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8
Q

What is the function of the 5’ cap?

A
  • Capping linked to transcription
  • Important structure for mRNA stability, production and function
  • The cap is a protein-binding element
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9
Q

What is Dystrophin?

A

Gene linked to Duchenne muscular dystrophy

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10
Q

What is the function of Conserved sequences in introns?

A
  • Intron and exon boundaries contain conserved sequences
  • Sequences define limits of exon and intron
  • Sequences recruit the splicing machinery required to remove the intron and join the exons
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11
Q

What is the structure of Conserved sequences in introns?

A
  • 5’ splice site
  • 3’ splice site
  • Branch site
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12
Q

What is the 2 step splicing of introns?

A
  • Step 1
    -cut at 5’ splice site
    -creation of bond between 5’ end of intron and branch site
  • Step 2
    -cut at 3’ splice site to release intron lariat
    -ligation of two exons
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13
Q

What is The spliceosome?

A
  • Enzymatic complex that catalyses the removal of introns
  • Requires ATP
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14
Q

What is the structure of The spliceosome?

A

*RNA-binding proteins
*ATPases
*GTPases
* SnRNPs

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15
Q

What are SnRNPs?

A

*Small nuclear ribonucleoprotein particles
*Stable RNA
*RNAs do not code for protein
*RNAs base-pair with conserved sequences in the intron
*Splicing is catalysed by the snRNAs

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16
Q

What is anti-Sm?

A
  • Anti-Sm antibodies react against the Sm proteins.
  • Anti-Sm antibodies are very rare unless you have the autoimmune disease systemic lupus erythematosus
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17
Q

What are the Mutations causing defects in splicing?

A

Spinal muscular atrophy
Retinitis Pigmentosa
Myotonic Dystrophy

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18
Q

What is Spinal muscular atrophy?

A

Most common genetic cause of infant mortality

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19
Q

What is Retinitis Pigmentosa?

A

Reduced visual capabilities and blindness

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20
Q

What is Myotonic Dystrophy?

A

A muscle wasting disease

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21
Q

What is the process of polydenylation?

A

Endonuclease cleavage
Addition of As by polyA polymerase
Bind CRSF and CstF.

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22
Q

What is CPSF?

A

Cleavage and polyadenylation specificity factor (CPSF)
binds AAUAAA

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23
Q

What is CstF?

A

Cleavage stimulatory factor (CstF)
binds G/U

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24
Q

What are the Functional significance of the polyA tail?

A
  • All mRNAs a 3’ poly(A) tail
  • Approx 250 nucleotides long
  • Bound by polyA-binding protein
  • Enhances export of RNA
  • Stabilises the 3’ end of the mRNA
  • Enhances translation of mRNA
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25
Q

What is RNA editing?

A

Nucleotide alterations which result in different or additional nucleotides in the mature RNA

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26
Q

Where does RNA editing occur?

A

mRNA
tRNA
ribosomal RNA (rRNA)

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27
Q

What are the Two classes of editing?

A

insertion/deletion
modification

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28
Q

What is N6-methyladenosine?

A

Regulated by writers, readers and erasers

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29
Q

Example of a writer.

A

Mettl3

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30
Q

Example of reader.

A

Hu-R
YTHDF1-3

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31
Q

Example of erasers.

A

FTP
AKLBH5

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32
Q

What is Enzymatic Deamination?

A

Change one nucleotide for another.

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33
Q

What is ApoB mRNA editing?

A

Editing carried out by the APOBEC-1 enzyme

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34
Q

What are the two types of ApoB mRNA editing?

A

Intestine (editing), ApoB-48
Liver (no editing), ApoB-100

35
Q

What is A to I editing in the Q/R Site of Glutamate Receptors?

A
  • L-glutamate major excitatory neurotransmitter
  • Editing yields decrease in Ca2+ permeability of channels containing the ‘R’ version
  • Editing carried out by ADAR2.
  • Mutations in the mouse ADAR2 gene
    lead to seizures, post-natal death,
    neurodegeneration in the hippocampus.
36
Q

What is ADAR2?

A

adenosine deaminase acting on RNA

37
Q

Why localise mRNA?

A
  • Localised protein synthesis
  • Generate cell polarity
  • Prevents expression in the wrong place
  • Promotes efficiency of subsequent protein targeting
  • Local control of translation
38
Q

How does RNA leave through the pore?

A

Cage like structure with a plug in the middle.
RNA is acidic and charged.
Pore is hydrophobic so need export factors.

39
Q

How does tRNA become a protein?

A

Initiation - Large subunit joins the small one.
Elongation - tRNA comes in with FTU docks tRNA in comes in at the A site.
-Peptide bonds form.
-Ribosome rotates and twists to pull mRNA through.
Termination - Peptide comes out the exit tunnel.
- Ribosome splits into subunits using energy.

40
Q

What is APE?

A

A - aminoacylated tRNA binding site
P - polypeptide chain site
E - exit site

41
Q

What is the kozak consensus sequence/

A

CC(A/G)CCAUG
Where the small subunit binds the CAP.

42
Q

What is eIF4E?

A

m7G binding

43
Q

What is eIF4G?

A

Binds eIF4E, A, 3 AND PABP

44
Q

What is eIF4A?

A

ATPase
RNA helicase

45
Q

What proteins are needed for mRNA circularisation?

A

eIF4E, G and PAB.

46
Q

What are the steps to translation initiation?

A
  1. Ribosome recycling
  2. eIF2 Ternary complex formation
  3. 43S complex formation
  4. Attachment to mRNA
  5. 5’ to 3’ scanning.
  6. Initiation codon recognition
  7. Hydrolysis of eIF2-bound GTP
  8. Subunit joining and factor displacement
  9. hydrolysis of GTP and release of eIF5B and eIF5B.
47
Q

What is eIF2B?

A

Governs levels of active GTP so initiation rate.
Activated by insulin.

48
Q

What is UTR?

A

Untranslated region

49
Q

How do iron levels affect mRNA translation?

A

Low iron, translational repression and mRNA stabilization.
High iron, translational activation and mRNA degradation.

50
Q

What is c-aconitase?

A

Interconverts citrate and isocitrate.

51
Q

What is the acceptor stem?

A

Where the amino acid is attached
They are reused.

52
Q

How are peptide bonds formed?

A

Preferred route is reversed.
Releases water.
Catalysed by ribosome.
Peptide always connected tRNA.
GTP docks onto the ribosome.

53
Q

What does eIF(-) stand for?

A

e eukaryotic
I initiation
F factor
(-) subunit

54
Q

What is the function of eIF?

A

Bind to the cap and allows ribosome to start translation.

55
Q

What is the function of eIF4G?

A

Opens up any structures between the cap and the AUG allows small subunits to scan along.

56
Q

What is the pre-initiation complex?

A

Scans to find the AUG.
Initiates hydrolysis of the eIFF2-boung GTP and phosphate release.

57
Q

What is eRF1?

A

Release factor causing the peptide release from tRNA.

58
Q

What is IP1/2?

A

Causes translational repression.
Lots of it transported into the cell to cope with low iron.

58
Q

What is IP1/2?

A

Causes translational repression.
Lots of it transported into the cell to cope with low iron.

59
Q

How does high iron lead too mRNA destabilization?

A

High levels of iron need storage.
Iron bind to IRP1.
IRP2 is degraded.
mRNA are destabilised

60
Q

What is the problem with IRP1?

A

When not bound to iron it can bind to mRNA.
RNA and Fe+ bind the same site, COMPETITIVE INHIBITION.

61
Q

Why is RNA degraded?

A
  • Damaged mRNA
  • Incorrectly transcribed/processed mRNA
  • Control gene expression
62
Q

What is casein mRNA?

A
  • mRNA increases
  • Half-life increases dramatically
  • Poly(A) tail length INCREASED
  • 3’ UTR of RNA binds proteins which aid in this stabilisation
63
Q

Why must mRNA become linear?

A

Closed loop must be broken before exonucleases can gain access

64
Q

How does mRNA become linear?

A

*lost cap or poly (A)
* brings ribosomes ending translation close to the AUG

65
Q

Examples of Decapping enzymes.

A

DCP1
DCP2

66
Q

Examples of endonucleases.

A

Argonaute
Swt1
Smg6

67
Q

Example of deadenylases.

A

Ccr/Not complex

68
Q

What is the exosome?

A
  • The exosome is the main 3’ to 5’ exonuclease in the cell
  • Involved in RNA turnover and processing
  • Multisubunit complex
  • The rest of the subunits function in RNA binding and unwinding
69
Q

What is XRN1?

A
  • 5’ to 3’ exonuclease
  • Involved in RNA turnover and processing
  • Also involved in transcription termination
  • Functions after decapping of the mRNA
70
Q

What is deadenylation-dependent decay?

A

This is a mechanism whereby all mRNAs gradually lose their poly(A) tails.

71
Q

What is the process of Deadenylation-independent decay?

A
  • Autoregulation- Rps28B binds its own message
  • Edc3 is one of several activators of decapping enzymes in the cell
  • Nucleases targeting specific substrates
72
Q

What is NMD?

A

Nonsense-Mediated Decay

73
Q

What leads to NMD?

A

Mistakes in the RNA are detected
RNA is targeted for degradation
Premature stop codons (PTCs) result from errors

74
Q

What is the mechanism?

A
  • In the first round of translation, EJCs are removed from the
    mRNA by the ribosome.
  • When ribosomes reach the PTC, an EJC remains
    downstream specific factors (UPF proteins) that are part of the EJC or are recruited to it interact with the RNA degradation machinery
75
Q

What is RNAi?

A

RNA interference

76
Q

What is siRNA?

A

small inhibitory RNA

77
Q

What is miRNA?

A

micro RNA

78
Q

What is RISC?

A

RNA induced silencing complex

79
Q

What is the structure of miRNA?

A

21-23 nucleotide RNAs

80
Q

What is the function of miRNA?

A

Perfect complimentary to target RNA.
Thought to be mainly viral defence mechanism.
Leads to the degradation of the target RNA.

81
Q

What is the structure of siRNA?

A

21-23 nucleotide RNAs

82
Q

What is the function of siRNA?

A

Imperfect complimentary to target RNA
Key gene regulatory mechanism in the cell
Leads to block in translation

Gene Expression (3 decks)

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