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CMB2001 > processing of mRNA transcripts > Flashcards

processing of mRNA transcripts Flashcards

1
Q

what are the two elements, added post-transcription, to eukaryotic mRNA

A

m7G Cap and poly A tails

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2
Q

are m7G Cap and poly Z tails encoded in the genome

A

No

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3
Q

what are the primary transcript processing methods

A

Capping, splicing, polyadenylation and editing

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4
Q

How is primary transcript processing coupled to transcription

A

RNA pol II CTD

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5
Q

What does RNA initially contain at 5’ end

A

triphosphate

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6
Q

what are the two steps in capping

A

1) GpppN structure
2) methylation

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7
Q

what process alters chemical behaviour of bases

A

methylation

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8
Q

what is the functions of m7G cap (4)

A
  • protects mRNA from degradation by 5’-3’ nucleases
  • Facilitates splicing
  • Facilitates export from the nucleus
  • Critical for translation of most mRNA
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9
Q

How are the m7G cap functions mediated

A

Through protein binding
CBP80/CBP20 in the nucleus and elF4 complex in cytoplasm

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10
Q

what does elF4 complex mediated in relation to m7G cap function

A

translation

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11
Q

what does CBP80/CBP20 mediated in relation to m7G cap function

A

processing/export

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12
Q

what is the order in which post-transcriptional modification

A

1) transcription and capping
2) Splicing and polyadenylation

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13
Q

which is generally longer in mammals, introns or exons

A

introns

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14
Q

Intron and exon boundaries contain conserved sequences, what and where are they ?

A

5’ splice site - exon 1
3’ splice site - exon 2
branch site - intron

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15
Q

what are the conserved sequences in exon/introns required for

A
  • define limits of exon and intron
  • Sequences recruit the splicing machinery required to remove the
    intron and join the exons
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16
Q

what type of reaction are the two steps of splicing of introns

A

2 - trans-esterification

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17
Q

what occurs in the two steps of intron splicing

A

1) cut at 5’ splice site & creation of bond between 5’ end of intron and branch site
2) cut at 3’ splice site to release intron lariat & ligation of two exons

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18
Q

what are spliceosomes

A

enzymatic complex that catalyses the removal of introns

19
Q

what do spliceosomes require

A

ATP

20
Q

what proteins make up spliceosome

A
  • large complexs containing RNA-binding proteins, ATPase, GTPase and several snRNPs
21
Q

what are snRNPs

A

Small nuclear ribonucleoprotein particles

22
Q

what do RNA do in terms of splicing

A

RNAs base-pair with conserved sequences in the intron (5’ splice site, branch point)
*Splicing is catalysed by the snRNAs

23
Q

what are the two types of snRNPs

A

Sm proteins and snRNAs

24
Q

what reacts against Sm proteins

A

Anti-Sm antibodies

25
Q

when would a person have anti-Sm antibodies

A

very rare unless a person has autoimmune disease systemic lupus erythematosus (SLE)

26
Q

what percentage of patients with SLE have anti-Sm antibodies

A

20%

27
Q

what catalyses the splicing mechanism

A

RNA

28
Q

explain Step 1 of the splicing mechanism using RNA

A

1) U1 & U2 binds to the 5’ and binding site respectively
2) U4,U6+U5 is then recruited, then U1+U4 is release
3) A bond is created between 5’ end of intron and the branch site
4) U6,U5 and U2 holds the free exon 1 together to the looped intron (still connected to exon2)

29
Q

what is the E complex

A

U1 at the 5’ splice site of the intron

30
Q

what is the A complex

A

U1 at the 5’ splice site of the intron
U2 is at the branch site

31
Q

What is the B complex

A

all RNA (U1,2,4,5,6) are joined along mRNA

32
Q

what is the B* complex

A

The B complex after U1+U4 is released

33
Q

What is the C complex

A

mRNA with U2,U5,U6 joined after intron lariat being formed and exon one being freed

34
Q

explain Step 2 of the splicing mechanism using RNA

A

The spliceosome cuts the mRNA at 3’, U2,6,5 hold the 2 exons together and allows them to bind
- lariat is released

35
Q

why is the number of proteins greater than genes in the genome

A

because there can be different organisations/arrangements of the exons after splicing

36
Q

what are the 3 conditions caused by mutations causing defects in splicing

A
  • Spinal muscular atrophy
  • Retinitis Pigmentosa
  • Myotonic Dystrophy
37
Q

how does polyadenylation occur

A
  • Endonuclease cleavage releases the mRNA from the DNA template when the a specific sequence is recognised
  • Addition of As by PolyA polymerase
38
Q

what is the recognised specific sequences which activates endonuclease cleavage

A
  • AAUAAA (conserved), 10-35 nucleotides upstream of polyA site
39
Q

what is downstream of poly(A) site

A

G/U or U rich tract

40
Q

what is upstream of AAUAAA

A

U-rich element ‘USE’

41
Q

what are the proteins required for polyadenylation, and where do they bind to

A
  • Cleavage and polyadenylation specificity factor (CPSF) = binds AAUAAA
  • Cleavage stimulatory factor (CstF) = binds G/U
    -Poly(A) polymerase
42
Q

what are the functional significance of the polyA tail

A
  • Enhances export of RNA
  • Stabilises the 3’ end of the mRNA
  • Enhances translation of mRNA
43
Q

give an example of a protein which doesnt have a poly A tail

A

histones

CMB2001 (11 decks)

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