B4 - Txn And Rna Processing

Question 1

If human genes are monocystronic, how is it organised in trypanosoma

Answer 1

Polycistronic (many genes same promoter)

Question 2

What sort of tandem repeated gene families can you find and give example

Answer 2

Housekeeping proteins or developmentally regulated proteins

Hsp or tubulin

Question 3

Is mature mrna likeneuk capped and poly A and how (unlike pre mrna)

Answer 3

Yes but different process - via transsplicing get cap

By polyadenylation then add tail

Question 4

How can you get evidence polycisteonic exist eg through calmodulin tandem gene copies or vsg221

Answer 4

Block maturation into individual transcripts via cycloleucine or uv light

Visualise whole transcript

To make sure no promoter in interfenic space = reporter genes with cloned dna sequences of interfenic regions and won’t see gfp being expressed if not promoter/monocystronic

Question 5

What are cis-regulatory elements interacting with trans-elements in euk expression. And what does rna pol need to interact with promoter element

Answer 5

Cis is sites like promoter

Trans are proteins/tf interacting

Rna pol needs initiation complexes to recognise promoter and bind

Question 6

What do the 3 rna pol transcribe usually

Answer 6

Rna pol 1 - RRNA
Pol II- pcg and U-snRna (uridine rich)
Pol III- trna

Question 7

Why is rna pol 1 different in trypanosoma

Answer 7

Encodes also some pcg eg parp and vsg

Question 8

What extra does pol III transcribe

Answer 8

U2 and 4 snrna

Question 9

How are the rna pol 1 promoters same for the transcripts eg vsg and parp

Answer 9

typical bipartite structure
Eg
Core promoter gc rich -45 to +20
Upstream gc rich region -180 to -107

Question 10

are the trans-activators known in trypanosoma

Answer 10

No

Question 11

How are promoters of normal pcg rna pol II different

Answer 11

They don’t have any!! Must have diff mech of transcription

Question 12

What does ctd (large subunit of rna pol 2) do in euk

Answer 12

Large subunit of rna pol II
Interacts with Tata binding proteins through heptapeptide repeats

Question 13

How is it diff in Tryp

Answer 13

phos

But not by tfIIH cak kinase domain because it doesn’t have one

Also no heptapeptide motif

Question 14

How is trans splicing diff to cis in euk

Answer 14

Cis-splicing is removing introns through forming a lariat via the spliceosome

Trans splicing is the addition of a 39bp splice leader sequence to 5’ of each spliced individual transcript from premrna polycistron

Question 15

What is done on 3’ at same time

Answer 15

Poly adenylation

Question 16

What is encoding these splice leader sequences , how many genes and where

Answer 16

Derived from SLrna 5’
Around 200 genes for SLrna somewhere not close to the mRNAs sometimes even on diff chromosomes to the transcripts
Transcribed through rna pol II

Question 17

How is it similar to cis

Answer 17

Because you’re rejoining 2 mrna transcripts to form mature rna

Polyadenylation via polyApolymerase

Also have u2,4 and 6 and sm proteins (but not 1 and 5 homologs)

Question 18

How many nucleotides are modified in the 5’ of slrna different to euk

Answer 18

4 (2 in euk)

Question 19

Which is the only known promoter of rna pol II

Answer 19

SLrna

Question 20

Which complex of polypeptides was found to bind tatabp on this promoter to induce txn

Answer 20

tSNAPc (t brucei)

Question 21

Is CTD important in this promoter

Answer 21

Yes (but not phos by tfIIH kinase cak

Question 22

What is methylated for translation on the cap nucleotides by metthltransferases

Answer 22

2’ o-ribose methylation

Question 23

RNA EDITING IN KT DNA OF MACXI CIRCLES

Answer 23

X

Question 24

What is meant by editing of the rna

Answer 24

After txn, (ptm) uridines are added or deleted from transcript to form mature mrna

These aren’t coded in the gene

Question 25

What gene in crithidia identified first for editing and what was added (maxicircle gene)

Answer 25

Cox subunit II (4 Us were added in 5’ end)

Question 26

What can U editing change

Answer 26

Stop codon, reading frame and start codons

Question 27

Give dramatic example

Answer 27

Cox subunit IiI 55% of nt are edited U residues in the last 2/3 of 3’ Inserted into polyA , codes a premature stop codon

Question 28

What do grna encdoed by minicircle a have on 5’ end of them

Answer 28

Anchor sequence complementary to the maxi circle dna 3’ of the editing site

Question 29

What do grna have on 3’ end

Answer 29

A poly U tail added post txn

Question 30

What adds and deletes Us at 3’ of cleaved fragment after cleaved by endonuclease

Answer 30

Tutase adds ExoUase removes

Question 31

What rejoins them using atp

Answer 31

Rna ligase

Question 32

What is the 20s macromolecular complex called for rna editing

Answer 32

Editosome

Question 33

What does it associate with

Answer 33

Kinetoplast rna editing KRE proteins Eg tutase, exoUase, ligase, helicase

Question 34

What frees the mrna strand from grna at end

Answer 34

Reh1 helicase

Question 35

What are the 2 subcomplexes found (3 proteins each) for addition or deletion

Answer 35

Addition complex - KrepA1, KREL 2, tutase Deletion Krepa2, KREL 1, exoUase

Question 36

What are krepA proteins (6 of them)

Answer 36

Those with ob folds for rna-protein interaction Unknown function

Question 37

What do the b barrel ob folds do

Answer 37

Ensure protein-rna interaction

Question 38

Which other multi protein complex found recently to interact with editosome

Answer 38

Mrb1 compelx

Question 39

The proteins contain which sorts of rna binding motifs

Answer 39

C2h2 zinc finger domain Or rrm rna BD (rna recog motif)

Question 40

What 2 proteins important in the mrb1 core subcomplex (2 subcomplexes)

Answer 40

Gap1 and 2

Question 41

What do they do / the core subcomplex

Answer 41

Initiate the editing by grna through stabilising it

Question 42

What is the other subcomplex called

Answer 42

TbRgg2 complex

Question 43

What is this for and how does rgg2 do this

Answer 43

3’ to 5’ mrna editing progression Rgg2 has high affinity rna recognition motif binding and binds rna

Question 44

How can you selectively block pol 2 and 3 to identify which is important at which point

Answer 44

A amanitin for pol 2 Tagetitoxin for pol 3

Question 45

Which sequence usually precedes the trans-splicing site in mrna processing (where SLrna is added)

Answer 45

Poly Y (pyrimidine chain)