B4 - Txn And Rna Processing Flashcards
If human genes are monocystronic, how is it organised in trypanosoma
Polycistronic (many genes same promoter)
What sort of tandem repeated gene families can you find and give example
Housekeeping proteins or developmentally regulated proteins
Hsp or tubulin
Is mature mrna likeneuk capped and poly A and how (unlike pre mrna)
Yes but different process - via transsplicing get cap
By polyadenylation then add tail
How can you get evidence polycisteonic exist eg through calmodulin tandem gene copies or vsg221
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
What are cis-regulatory elements interacting with trans-elements in euk expression. And what does rna pol need to interact with promoter element
Cis is sites like promoter
Trans are proteins/tf interacting
Rna pol needs initiation complexes to recognise promoter and bind
What do the 3 rna pol transcribe usually
Rna pol 1 - RRNA
Pol II- pcg and U-snRna (uridine rich)
Pol III- trna
Why is rna pol 1 different in trypanosoma
Encodes also some pcg eg parp and vsg
What extra does pol III transcribe
U2 and 4 snrna
How are the rna pol 1 promoters same for the transcripts eg vsg and parp
typical bipartite structure
Eg
Core promoter gc rich -45 to +20
Upstream gc rich region -180 to -107
are the trans-activators known in trypanosoma
No
How are promoters of normal pcg rna pol II different
They don’t have any!! Must have diff mech of transcription
What does ctd (large subunit of rna pol 2) do in euk
Large subunit of rna pol II
Interacts with Tata binding proteins through heptapeptide repeats
How is it diff in Tryp
phos
But not by tfIIH cak kinase domain because it doesn’t have one
Also no heptapeptide motif
How is trans splicing diff to cis in euk
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
What is done on 3’ at same time
Poly adenylation
What is encoding these splice leader sequences , how many genes and where
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
How is it similar to cis
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)
How many nucleotides are modified in the 5’ of slrna different to euk
4 (2 in euk)
Which is the only known promoter of rna pol II
SLrna
Which complex of polypeptides was found to bind tatabp on this promoter to induce txn
tSNAPc (t brucei)
Is CTD important in this promoter
Yes (but not phos by tfIIH kinase cak
What is methylated for translation on the cap nucleotides by metthltransferases
2’ o-ribose methylation
RNA EDITING IN KT DNA OF MACXI CIRCLES
X
What is meant by editing of the rna
After txn, (ptm) uridines are added or deleted from transcript to form mature mrna
These aren’t coded in the gene
What gene in crithidia identified first for editing and what was added (maxicircle gene)
Cox subunit II (4 Us were added in 5’ end)
What can U editing change
Stop codon, reading frame and start codons
Give dramatic example
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
What do grna encdoed by minicircle a have on 5’ end of them
Anchor sequence complementary to the maxi circle dna 3’ of the editing site
What do grna have on 3’ end
A poly U tail added post txn
What adds and deletes Us at 3’ of cleaved fragment after cleaved by endonuclease
Tutase adds
ExoUase removes
What rejoins them using atp
Rna ligase
What is the 20s macromolecular complex called for rna editing
Editosome
What does it associate with
Kinetoplast rna editing KRE proteins
Eg tutase, exoUase, ligase, helicase
What frees the mrna strand from grna at end
Reh1 helicase
What are the 2 subcomplexes found (3 proteins each) for addition or deletion
Addition complex -
KrepA1, KREL 2, tutase
Deletion
Krepa2, KREL 1, exoUase
What are krepA proteins (6 of them)
Those with ob folds for rna-protein interaction
Unknown function
What do the b barrel ob folds do
Ensure protein-rna interaction
Which other multi protein complex found recently to interact with editosome
Mrb1 compelx
The proteins contain which sorts of rna binding motifs
C2h2 zinc finger domain
Or rrm rna BD (rna recog motif)
What 2 proteins important in the mrb1 core subcomplex (2 subcomplexes)
Gap1 and 2
What do they do / the core subcomplex
Initiate the editing by grna through stabilising it
What is the other subcomplex called
TbRgg2 complex
What is this for and how does rgg2 do this
3’ to 5’ mrna editing progression
Rgg2 has high affinity rna recognition motif binding and binds rna
How can you selectively block pol 2 and 3 to identify which is important at which point
A amanitin for pol 2
Tagetitoxin for pol 3
Which sequence usually precedes the trans-splicing site in mrna processing (where SLrna is added)
Poly Y (pyrimidine chain)
