Lecture 5/6: Regulation of the transcriptiome Flashcards
(27 cards)
What are two modifications made to the ends of eukaryotic mRNA
capping 5’ end and polyadenylation on the 3’ end
What is a key step in transcription initiation by RNA polymerase II
Phosphorylation of the RNA polymerase II tail, also called the C-terminal domain (CTD)
Why is CTD phosphorylation key in initiating transcription
- helps dissociate RNA polymerase II from other proteins present at the start of transcription
- allows a new set of proteins that function in transcription elongation and RNA processing to associate with the RNA polymerase tail
What do red and green indicate on a DNA microarray
green= little RNA
red= lots of RNA
black= middle
Describe how DNA microarrays were used to analyze AML
- primary AML cells= from patients
- confirmed that model cell for AML, undifferentiated HL-60 exhibited same behaviour in the transcription as primary cells
- Treated HL-60 with different chemicals to find one that would modify the transcription such that it is the same as the differentiated neutrophils (normal)
- used blue to indicate little RNA and red to indicate lots
- many genes showed that AML vs normal cells had opposite expression; this does not mean that every single gene is opposite, only that those that were tested presented that way
What is IC50
the concentration of a compound required to inhibit cell proliferation by 50%
lower concentration (lower IC50) required= stronger and more effective compund
What are the functions of RNA capping
1) Helps in RNA processing and export from the nucleus
2) Important role in translation of mRNAs in the cytosol
3) protects mRNA from degradation
What is RNA capping
addition of a modified guanine nucleotide to the 5’ end of pre-mRNA using 3 enzymes
Cap bound using a cap-binding complex (CBC)
What are coding and noncoding sequences in eukaryotic genes called?
coding= exons
noncoding=introns
introns are removed via RNA splicing
Describe drosphila sex determination ratios
chromosomes : autosomal sets
X:A= o.5=male
X:A= 1.0=female
below o.5= meta male
between 0.5-1.0= intersex
above 1.0= meta female
What genes are involved in drosophila sex determination and what do they do
sex-lethal: splicing repressor
transformer: splicing activator
double sex: reglates gene expression (represses male/female gene expression)
How to determine drosophilla sex
1) count X chromosomes
2) ignore Y chromosomes
3) count # autosomal sets (one set = 3 chromosomes)
What is CstF
Cleavage stimulating factor, used in 3’ polyadenylation
What is CPSF
cleavage and polyadenylation specificity factor, used in 3’ polyadenylation
Describe polyadenylation
- RNA cleaved
- transcrition terminates
- Poly-A-[olymerase (PAP) adds around 200 A nucleotides to the 3’ end of RNA to aid in RNA export, translation, and mRNA stability
not genome encoded
Describe alternative splicing in drosiphila males
no regulation at sxl, tra, or dsx. dsx protein represses female gene expression
Describe alternative splicing in drosophila females
splicing regulated at all levels.
- sxl protein is functional and represses the splicing of sxl and tra
- tra not spliced, functional tra protein produced
- functional tra protein and Tra2 bind to activate the splicing of dsx
- dsx protein represses male differentiation genes leading to female development
What degrades improperly processed mRNAs
exosomes
What are two initiation factors and where do they bind
eIF4E bound at 5’ cap
eIF4G biound at Poly-A-Binding protein
What are the functions of the eIFs
OCCURS IN CYTOSOL; part of post-TRANSCRIPTIONAL regulation
-recruit small ribosomal complex which will initiate translation at first AUG downstream of 5’ cap
- ensures that both ends of mRNA are intact
What is the function of the EJC
OCCURS IN CYTOSOL; part of post-TRANSCRIPTIONAL regulation
exon junction complex
- stimulates translation to ensure proper splicing
- displaced (ran thru) by the moving ribosome in normal splicing so that none remain by time the stop codon is reached
- indicator of abo=normal splicing, as they remain on the mRNA when the ribosome reaches the stop codon
What is the function of nonsense-mediated mRNA decay
- may have played an important role in th evolution of eukaryoted by allowing for selection of DNA rearragements to produce full-length proteins
-important in immune cells for antibody production
What is the prokaryothic method of quality control for broken mRNAs
- ribosomes stall on broken or incomplete mRNAs and do NOT release
- a secial RNA called tmRNA carries an alanine AA and acts as both tRNA and mRNA
- releases mRNA once tmRNA binds to A site
- alanine added onto the polypeptide from tmRNA
- ribosome translates 10 codons, forming an AA tag that can be recognized by proteases for degradation
What degrades broken RNAs in prokaryotes
exonucleases
