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Biochemistry > RNA Splicing > Flashcards

RNA Splicing Flashcards

0
Q

R

A

Purine

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1
Q

Exon

A

Any region retained in a mature mRNA

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2
Q

Y

A

Pyrimidine

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3
Q

N

A

Any (purine or pyrimidine)

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4
Q

Key recognition in humans

A

5’…(C/A)G-GU(A/G)AGU………YNYURAY………(Y11)NCAG-G…3’

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5
Q

Transesterification

A

The process of exchanging the organic group R” of an ester with the organic group R’ of an alcohol

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6
Q

Nuclear pre-mRNA splicing

A
  1. OH-2’ attaches to P on the 5’exon side
  2. OH-3’ at the end of 5’exon attacks phosphate at the beginning of the 3’-exon
  3. Forms intron lariat and spliced exon bonded (mature mRNA)
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7
Q

Spliceosome roles

A
  1. Recognize 5’ splice site and branch site
  2. Bring these sites together
  3. Help catalyze RNA cleavage and joining
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8
Q

Assembling the Spliceosome

A
  1. snRNA U1 recognizes 5’ splice site
  2. U2AF bridges intron-exon boundary
  3. Interaction between U2AF and BBP allows BBP to bind to branch site
  4. Displacement of BBP by snRNP U2 (with help from U2AF) results in formation of A complex
  5. U2 causes A to pop making it more available to bind
  6. Rearrangement of A complex mediated by tri-snRNP particle (U4-U5-U6) forms B complex. U2AF leaves
  7. Segment of RNA in front of A is bent
  8. U1 is replaced by U6 completing Spliceosome.
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9
Q

U1 sequence

A

CAUUCA

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10
Q

U6

A

GAGACA

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11
Q

U2 sequence

A

AUGAUG

binds to A site

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12
Q

Canonical pathway splicing

A
  1. U4 is displaced from the complex
  2. Allows U2 and U6 to interact forming the active site
  3. Transesterification #1: 5’ splice site to branch site
  4. Transesterification #2: 5’ and 3’ splice sites brought together by U5.
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13
Q

Group II self-splicing introns

A

Not enzymes

  • mechanisms same as canonical
    1. “A” residue attacks phosphodiester bond at 5’ slice site
    2. 3’-OH attacks 3’ splice site
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14
Q

Group 1 self-splicing introns

A

Not enzymes - uses free G nucleotide or nucleoside

  1. 3’-OH attacks 5’ splice site
  2. New 3’-OH attacks 3’ splice site
  3. Linear intron released
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15
Q

Potential splice site recognition errors

A
  1. Exon skipping
  2. Pseudo splice-site selection - somewhere on exon has a sequence so similar to intro sequence that machinery confuses it for an intron
16
Q

How can splice site accuracy be enhanced

A

Cotranscriptional loading

17
Q

Cotranscriptional loading

A

During transcription - pol tail prepares premRNA for splicing by adding components

18
Q

Another mechanism for increasing splice site accuracy

A

Serine-argine (SR) rich proteins bind exonic splicing enhancers

SR-ESE interactions recruits U2AF to 3’ splice site and U1 snRNP to the 5’ splice site

Function of SR is to show where splice starts and ends (not always on exons)

19
Q

Trans-splicing

A

2 exons from different RNA molecules are spliced together

Y-shaped rather than lariat

Same machinery minus U1

20
Q

Alternative splicing

A

Generates diversity of gene expression

Picks and chooses which exon/intron gets spliced out

Increases genetic diversity without extending DNA

21
Q

snRNP steric hindrance

A

Depending on which protein (U1 or U2) you get different cuttings in DNA

U1 binds first U2 can’t bind
U2 binds first U1 can’t bind

U1 and U2 always bind in pairs

22
Q

What helps ensure U1 and U2 to bind in pairs

A

Their tails

23
Q

Repressive mediated alternative splicing

A
  1. Some splicing sites had a repressor site to regulate splicing.
  2. If a repressor protein is bounded to the repressor the splicing machinery cannot bind to splicing site
  3. mRNA does not get spliced (at least at that particular site)
24
Q

Activator-mediated alternative splicing

A
  1. In order to splice a certain region of the mRNA an activator must bind to the splicing enhancer site
  2. Once we have a splicing machine and an activator the specific site can be spliced out.
  3. Without the activator, the site will remain unspliced in the final mRNA product

Biochemistry (10 decks)

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