L7-8: Post transcriptional control of gene expression I

Question 1

prokaryotic txn and translation

Answer 1

one compartment
coupled

Question 2

eukaryotic txn and translation

Answer 2

separate membrane compartments
txn in nucleus moved to cyto for tln
uncoupled

Question 3

stages of eukaryotic gene expression

Answer 3

1. txn control
2. RNA processing control
3. tln control
4. protein activity control

reg at each stage

Question 4

eukaryotic mRNA structure

Answer 4

1. m7 Gcap structure
2. start codon
3. coding region
4. stop codon
   5
5. polyA tail

start codon > AUG
poly A = ~250 A’s

Question 5

m7 Gcap and polyA tail

Answer 5

specific to euk mRNAs
post-txn added
encoded in genome

Question 6

capping

Answer 6

coupled to txn via RNA pol II
C-Terminal Domain as binding site

Question 7

intron conserved sequences

function?

Answer 7

define limits of exon/ intron
recruit splicing machinery for intron removal / exon joining

Question 8

2-step intron splicing

2 step esterification

Answer 8

1. cut at 5’ splice site
2. cut at 3’ splice site

Question 9

5’ m7 Gcap structure

Answer 9

all RNA pol II RNAs
initially contains triphosphate at 5’
sugar phosphate bond

Question 10

5’ m7Gcap synthesis

Answer 10

1. GpppN structure
2. methylation

methylation alters base chemical behaviour

Question 11

m7 Gcap functions

Answer 11

protects RNA from degardation by 5’-3’ nucleases
facilitates splicing/ export from nucleus
critical for mRNA translation
protein-binding element

important for mRNA production/ stability/ function

Question 12

M7 Gcap function mediation

Answer 12

CBP80/20 in nucleus
elf4 in cytoplasm

via protein binding

Question 13

alternative splicing

Answer 13

1. txn and capping// introns cut out for exon joining> forms pre-mRNA
2. splicing and polyadenylation (non-coding removed) > forms mRNA

Question 14

Intron length

Answer 14

no. length of introns vary per gene
usually 80-10,000 nt long

intron length> exon length

Question 15

dystrophin

Answer 15

gene linked to Duchenne muscular dystrophy
1 intron
21000 nt

Question 16

intron conserved sequence components

Answer 16

5’ splice site
3’ splice site
branch site

Question 17

cut at 5’/3’ splice site in 2 step esterification

Answer 17

5’> bond formed between 5’ end of intron and branch site
3’> release intron lariat/ ligation of 2 exons

Question 18

spliceosome

Answer 18

enzymatic complex that catalyzes intron removal

complex of >200 proteins assembling onto each intron

(proteins: RNA-binding/ ATPases/ GTPases/ snRNPs)

Question 19

snRNPs

Answer 19

small nuclear ribonucleo-protein particles
stable RNA-protein complexes in nucleus
not RNA coding
catalyse splicing
Sm ring (binding site of RNA)

U1/2/4/5/6 in spliceosome

Question 20

snRNP base-pairing w conserved sequences in intron

Answer 20

U1> 5’splice
U2> branch

Question 21

SLE

systemic lupus erytematosus

Answer 21

~20% have anti-Sm

anti-Sm Ig react against Sm proteins

Question 22

txn and splicing

Answer 22

functionally coupled
yeast U1/U2 pair to pre-mRNA

Question 23

no. conserved sequences in splicing

Answer 23

3

Question 24

no. introns in S.cerevisiae vs C.elegans

Answer 24

minimal splicing/ introns in S.cerevisiae
many introns in human genome/ C.elegans

Question 25

alternative splicing function

Answer 25

key for multiple protein isoforms from one gene

Question 26

types of alternative splicing

Answer 26

exon skipping
intron retention
mutually exclusive exons
alternative 5’/3’ splice site

during embryonic development

Question 27

activators

in regulation of cis-acting sequences in pre-mRNA

Answer 27

bind to intronic/ exonic splicing enhancers
ISE/ESE

intronic/exonic splicing enhancers

Question 28

repressors

in regulation of cis-acting sequences in pre-mRNA

Answer 28

bind to intronic/ exonic splicing silencers

Question 29

mutations causing splicing defects

Answer 29

spinal muscular atrophy (infant mortality)
retinitis pigmentosa (blindness/ vision defects)
myotonic dystrophy (muscle-wasting)

Question 30

polyadenylation

2 stages

Answer 30

1. endonuclease cleavage
2. addition of As by polyA polymerase

Question 31

conserved sequence for polyadenylation

other features of polyadenylated mRNA?

Answer 31

AAUAAA
10-35nt up of poly(A) site

G/U or U rich tract downstream of poly(A)/ USE

proteins bind to these sequences

Question 32

USE

Answer 32

U-rich upstream element

Question 33

protein binding AAUAAA

Answer 33

Cleavage and polyadenylation specificity factor

CPSF

cleavage factors I/II

Question 34

protein binding G/U

Answer 34

cleavage stimulatory factor

CstF

Question 35

poly(A) tail

no. nt? location?

Answer 35

250 nt
3’ on all mRNA
bound by poly(A) binding protein

Question 36

poly(A) tail function

Answer 36

^RNA export/ mRNA tln
protein-binding element
stabilizes 3’ end of mRNA

Question 37

2 RNA editing classes

where does it occur

Answer 37

insertion/ deletion
modification

mRNA/tRNA/rRNA

Question 38

RNA editing in medicine/ development

Answer 38

atherosclerosis
brain function improvement
drosophilia development
parasites (trypanosoma leishmania/ trypanosoma)

Question 39

base modification

functions

Answer 39

middle group addition
aids protein-binding
flipping purines/ pyrimidines

marked nucleotide/ altered identity

Question 40

effects of mRNA editing

Answer 40

U-insertion> start-codons/ stop-codons
nt insertion> new ORF’s
change in encoded aa/ splice site by base conversion
stop codons removed by base conversions

C> U changes

Question 41

enzymatic deamination

Answer 41

adenosine> inosine
cytosine> uracil

inosine recognised as guanosine

Question 42

cytidine deamination

apoB pre-mRNA editing

Answer 42

• APOBEC-1 enzyme carrying out cholesterol control/ Cancer development/ viral replication inhibition
• 2 forms(48/100) circulate in blood w different functions

100> liver w/o editing

Question 43

2 forms of APOBEC-1

Answer 43

48> intestine w editing
100> liver w/o editing/ long form component of LDL (atherosclerosis linked)

Question 44

L-glutamate

Answer 44

major excitatory neurotransmitter

Question 45

A> I editing in Q/R site of glutamate

Answer 45

decreases Ca2+ permeability of ‘R’ channels
carried out by ADAR2

Question 46

ADAR2

Answer 46

Adenosine Deaminase on RNA

Question 47

ADAR2 mutations

Answer 47

seizures/ post-natal death/ neurodegeneration in hippocampus

Question 48

nucleotide modification

Answer 48

nt modification after txn
results in changes in protein binding> changes in RNA properties/ sequences

editing on ncRNAs also> essential for function

Question 49

mRNA export from nucleus to cytoplasm

Answer 49

nucleopores in nucleoplasm
via nuc basket
cyto/proximal filaments
central transporter
cyto/inner spoke/ nucleoplasmic ring

different pathways mediate export of different RNAs

Question 50

mRNA localisation fucntion

Answer 50

localised protein synthesis
cell polarity generation
prevents expression in wrong place
^protein targetting efficiency
tln control

Question 51

dendritic mRNA localisation

Answer 51

tln local for synaptic plasticity/ spine morphogenesis

Question 52

mRNA localisation by diffusion

Answer 52

mRNA’s freely diffuse in cytoplasm/ locally entrapped by anchor proteins

Question 53

active transport- based localisation

Answer 53

• mRNA recognized by specific trans-acting factors in nucleus
• cytoplasmic factors ensure transport along polarized cytoskeleton