eukaryotic transcription

Question 1

transcription of DNA to RNA

Answer 1

• RNA pol needs general TF to recruit it and bind DNA binding site in promoter
• TF bind enhancers in distal location
• mediator links enhancer to RNA pol and stabilizes initiation complex, chromatin opens up
• RNA starts initiation, but then pauses due to negative EF
• need P-TEFb, which is recruited by Myc
• P-TEFb phosphorylates neg EF and RNA pol II to begin elongation

Question 2

“modulator” transcription factors

Answer 2

• TF have two domains: DNA binding and enhance/repress
• can separate and make new proteins
• in cancer: activator domain moves next to PBX 1 –> leukemia
• T cell acute lymphocyte leukemia: T cell enhancer moves next to HOX gene –> too much HOX protein
• Burkitt lymphoma: enhancer translocates next to MYC, an oncogene with many targets to gene expression, get too much MYC and too much expression

Question 3

TF binding domain

Answer 3

• alpha helix of TF binds major groove of DNA
• each TF has specific AAs in side chains for specificity
• once it binds, recruits transcription machinery or chromatin modifiers

Question 4

defects in transcription

Answer 4

• blocking differentiation –> cells remain immature and continue dividing –> cancer
• can miss express a cell in wrong location
• oncogenes and tumor suppressors are TFs

Question 5

chromatin modification

Answer 5

• chromatin must loosen to RNA pol to work

- can modify histones, nucleosomes or DNA

Question 6

histone modification

Answer 6

-acetylate, deacetylate, methylate, phosphorylate
“writers” add
“erasers” take off
“readers”
drugs target these to inhibit modifications or so readers can bind modified histones

Question 7

RNA processing

Answer 7

-5’ guanine cap is 5-5 link
-3’ poly A tail - made w/o template
these protect, facilitate export and help start translation
-also involves splicing

Question 8

introns

Answer 8

• big genes allow for possibility of recombination –> genetic diversity
• alternative splicing allows different isoforms of same protein that are specific for tissue, time and function
• help with regulation

Question 9

splicing mechanism

Answer 9

-cis sequences in donor and acceptor site. take out RNA in between

Question 10

splicing disease

Answer 10

• Hutchinson Gilfor Progeria
• C–> U makes the sequence a splice site
• get truncated protein that cannot remove Lamin A from cell membrane

Question 11

splicing treatment

Answer 11

• Spinal muscular dystrophy SMN1 mutation causes faulty splicing for SNRPs
• drugs prevent splicing repressor from working so the RNA transcript includes all exons and get functioning protein
• for Alzheimers: TDP43 aggregates incytoplasm of neurons and sequesters RNA
• treat using the DBR1 mutant of lariot debranching enzyme that cannot degrade lariot
• lariots can sequester excess TDP43, protecting neurons from TDP43 aggregation

Question 12

non coding RNA gene regulation

Answer 12

siRNA - cleave target mRNA
miRNA - base pair with mRNA and prevent translation or cause degradation
piRNA - prevents transposition into germline
long ncRNA - form regulatory complexes. upregulated in cancer and people at risk for CVD and stroke

Question 13

RNA seq

Answer 13

• measures type and amount of RNA in all cells

- ENCODE looks to map all epigenetic sites to map all the enhancers, splice sequences, etc.

Question 14

EQTL

Answer 14

• expression quantitative trait loci

- look at RNA expression in certain tissues to diagnose disease. is a “gene marker.” then find cause

Question 15

GWAS studies

Answer 15

• genome wide studies
• shows many disease linked to gene transcription and not protein mutation
• 90% link to non-coding RNA

Question 16

nobel prize

Answer 16

showing that differentiated cells may be turned back to stem cells with just a few TF
-then fixed, reinserted into patient and re-mature