post transcriptional mod

Question 1

addition of a 5’ cap
what is needed
and what happens

Answer 1

7-methyl-guanosine
caps as soon as RNA leaves the exit hole in pol 2
cap enzyme bound to RNA pol tail (p-ser-5)
gamma phosphate removed by RNA triphosphatase on RNA
guanylytransferase removes Y and beta of GTP and adds GMP to RNA
methyltransferase adds methyl group to guanine

Question 2

RNA triphosphatase

Answer 2

removes gamma phosphate off 5’

Question 3

guanylytransferase

Answer 3

removes gamma and beta phosphates off GTP and adds to RNA

Question 4

methyltransferase

Answer 4

adds methyl group to guanine

Question 5

what does the cap do

3 points

Answer 5

helps distinguish mRNA from RNA
helps mRNA be properly processed and exported to nucleus
protects against degradation

Question 6

3’ polyadenylation

what happens

Answer 6

adding of adenosine to 3’ end - 200bp
signals for this encoded in DNA
proteins bind to the poly a tail to help stabilise the RNA

Question 7

RNA splicing what is required

Answer 7

conserved sequence motifs indicating intron . exon boundries
starts GU
AG ends
branch point

Question 8

mechanism of RNA splicing

2 steps : step 1

Answer 8

introns removed by 2 transesterification reactions
step 1: -OH of A (branch point) acts as a nucleophile and attacks the 5’ splice site
intron folds back on itself
phosphodiester bond between A and G (from GU)

Question 9

mechanism of RNA splicing

step 2

Answer 9

cleavage of the 3’ splice site and joining exons
3’ -OH at start site acts as a nucleophile and attacks the AG end site
exons join and intron forms a lariet

Question 10

how is the splicing formed (ie enzyme formation etc)

Answer 10

co-transcriptional loading
RNA pol carries proteins involved in splicing
splicing machinery assembles as the gene is being transcribed

Question 11

spliceosome what is it made of

Answer 11

both RNA and proteins
U1, 2, 4, 5, 6
each RNA is complexed with 6-10 proteins to form snRNPs

Question 12

snRNAs

snRNPs

Answer 12

small nuclear RNAs

small nuclear RNPs

Question 13

what happens in the spliceosome

Answer 13

U1 binds to splice site
then BBP (branch point binding protein binds)
as well as U2AF35/65 which are (u2 auxiliary factors) helps position BBP

Question 14

then after u2 and u1 binds what happens

Answer 14

U2 binds by displacing BBP and is aided by U2AFs
which allows the RNA to bend
and calls in U4, 5 AND 6 while the U2AFs are displaced

Question 15

3 classes of RNA splicing

and what branch site do they use

Answer 15

nuclear pre-mRNA splicing (most common) branch site a
group 2 self splicing, branch site a
group 1 self splicing, branch site G

Question 16

what are SR proteins and why are they important

Answer 16

they are serine and arginine rich proteins
important for splice site slection
bnd to sequence called exonic splicing enhancer ESEs
helps recruit machinery

Question 17

alternate splicing what was the first gene found

Answer 17

calcitonin

Question 18

editing reaction

deaminase reaction ADAR what happens

Answer 18

glutamate receptor gets a A to I (inosine)
catalysed by adenosine deaminase action on RNA (ADAR)
alters the ca2+ permeability of the cell

Question 19

if ADAR was knocked out in mice what will happen

Answer 19

causes epilepsy and death

because it suppresses interferon signalling and to block premuture apoptosis

Question 20

transport of RNA through nuclear pores

Answer 20

REF associate with the spliceosome at the splicing junction
and remains attached
TAP and MEX bind to REF
allows transport of RNA through pore