final (exam 21,22) Flashcards
(70 cards)
1
Q
similarities between pro and euk transcriptional regulation
A
- mediated by DNA-binding proteins
- can be controlled by activators
- regulation via: recruitment of RNAP (pol 2 for mRNA in euk), open complex formation, and promoter escape
2
Q
heterochromatin
A
- tightly condensed DNA that is transcriptionally “silenced”
- 10% of chromatin: telomeres, centromeres, inactivated X chrom
- DNA is often methylated at GpC dntd repeats
3
Q
euchromatin
A
- less tighly condensed
- does not equate to active transcription, can POTENTIALLY be expressed
- histones prob acetylated (loosens chromatin structure, DNA becomes more accessible to proteins)
4
Q
how can enzymes change histone and chromatin structure?
A
by modifying histone tails through methylation, phosphorylation, ubiquination, and acetylation
each modification has a specific function- histone code
5
Q
HATs vs HDACs
A
HATS: heterochromatin –> euchromatin, bromodomains with histones
DHACs: euchromatin –> heterochromatin chromodomains
6
Q
H2AZ AND H3.3
A
replace H2 and H3 in euchromatin via alternate histone chaperones and maintain more “open” chromatin state
(active genes more often because loose)
7
Q
enhancers
A
activate transcription selectively
can be >10,000 bp upstream or downstream of a core promoter (yeast upstream activator sequences)
8
Q
how long are yeast upstream activator sequences usually?
A
< 200 bp upstream
9
Q
where do general transcription factor bind?
A
at all promoters
10
Q
activators
A
- bind to enhancers
- do not interact directly with RNAP. instead, recruit:
- proteins to modify chromatin structure and/or interact with mediator and/or GTP
-nucleosome remodelers that expose regulatory sequences like core promoters
-histone modifying enzymes (like HDACs) to loosen chromatin and mediator
-TF2D, GTF, and Pol 2 to exposed promoter
11
Q
what are activators made up of?
A
AD (activation domains)
BD (binding domains)
12
Q
4 common DNA binding motifs in eukaryotic activators (TF)
A
- homeodomain
- zinc finger
- leucine zipper
- helix-loop-helix
13
Q
number of regulatory sequences outside core promoter for euk and prok genes?
A
euk: 5+
prok: 1 or 2
14
Q
ChIP
A
- chromatin immunoprecipitation
- to see where particular proteins are bound on DNA
- detect: binding sites and distribution of TF; modification to histones that influence chromatin structure and gene expression
- fragments: 150-900 bp
15
Q
ChIP-exo
A
- can identify protein-DNA interactions with near bp precision (more exact)
- can remove DNA that is not actually bound by protein (chops up around the protein –> bp accurate)
16
Q
ChEC-seq
A
- add MNase to protein via flexible linker
- add Ca2+ (activates MNase)
- cleaves
- get small fragments of DNA
(MNase cuts area around YFP)
17
Q
liquid/liquid phase separation
what is it caused by?
A
- caused by intrinsically disordered regions of proteins
- increases concentration of TF
- can regulate whether they form or not
key roles in GENE EXPRESSION
18
Q
repression mechanisms
A
compete for binding to a regulatory region
prevent:
activator from interacting with mediator or other coactivators
coactivator from interacting with GTFs
19
Q
CpG length
A
- often methylated
- 1000-2000 b long
- several hundred bp upstream or downstream of transcription start site
20
Q
what causes Fragile X mental Retardation (FMR)
A
methylation of an expanded CGG repeated
-most common inherited cause of retardation
-normal: 6-50 copies
-FMR: >200
-1/1500 males
21
Q
how many UASgal do galactose utilization genes have?
how big?
A
1 or more (17bp enhancer motifs)
22
Q
Gal80p vs galactose absence:
A
Gal80p: Gal4p activates transcription
Galactose: Gal80p blocks Gal4P AD from interacting with TF2D
23
Q
what does the mediator stimulate?
A
TF2H: opens complex and promoter escape
24
Q
how can DNA methylation be detected?
A
by bi-sulfite sequencing
C is not converted to U (as it is supposed t)o
25
what does CTCF bind? what does that do?
insulator DNA
blocks activators from insulator DNA
blocks spread of histone or DNA modification (HATs/HDACs/methylases)
26
how many zinc finger domains does CTCF have?
11
27
CTCF bound at insulator probably acts by......................
creating loops in DNA
28
DNA methylation state is .......... with each round of DNA replication
inherited
29
maintenance methylases
recognize hemi-methylated DNA and copy the pattern/ modification to daughter strand without it
30
imprinting
-depending on the gene, either copy from mom or dad is
**epigenetically silenced**
-ususally due to DNA methylation during egg/sperm formation
-epigenetic tags on imprinted genes usually stay put for the life of the organism but are reset during egg/sperm formation
-regardless of whether they have mutations, certain genes are always silenced in the egg and others are always silenced in the sperm
31
imprinting: allele-specific silencing
parent with CTCF bound to insulator: expression off
parent with methylated insulator: CTCF cant bind, copy is active
32
Prader-Willi syndrome
-only paternal copy of gene expressed
-mental disabilities, behavioral issues, and always hungry
33
Angelman syndrome
-only maternal copy of gene expressed
-intellectual and development disability
-jerky movement and hand flapping
-frequent laughter and smiling
-unusually happy disposition
DUE TO MISSING/DEFECTIVE UBIQUITIN LIGASE
34
what is IGF2 involved in?
fetal growth and development
35
how many genes does genomic imprinting affect? what type?
100 genes
growth and development
36
genomic imprinting model
conflict of interest between parents over the resources allocated to the fetus by the mother during intrauterine gestation
37
JAK-STAT pathway
1. cytokine binds to JAK receptor
2. receptors dimerize and autophosphorylate
3. STAT binds, dimerizes, and autophosphorylates
4. STAT-P dimer enters nucleus and binds enhancer to activate immune system
38
how could you inhibit the immune system?
inhibiting STAT, which binds enhancer and activates transcription of genes involved in immune system function
39
how can a protein sometimes be bound to plasma membrane and other times be secreted into the plasma?
depending on how it was spliced --> alternate splicing
40
what is Gcn2? what does it do?
stress sensor
phosphorylates elF2 ---> elF2B is sequestered so it cannot bind to tRNA with Met ---> translation is blocked
shuts down energy hogs to turn on genes you need
41
Gcn4
-TF for amino acid synthesis in yeast
-hops on cap, rides along until find a good enough Kozak sequence (5th AUG
42
Gcn4 when:
there are lots of amino acids
vs
amino acid starvation
**abundant**: translation initiated on short decoy uORF, no Gcn4 produced
**starvation**: phosprhotylate elF2, ignore decoy uORF, initiate on 5th AUG (best for elF2), produce Gcn4
43
what does high production of Gcn4 do?
-increase amino acid synthesis
-lower ribosome protein synthesis
-increase stress response genes
44
what do IRP/IRE do when they are in 3'UTR?
regulate mRNA stability
45
what is transferrin?
when is it needed?
-iron chelator that helps scavenge extracellular ion
-needed when iron is low
-not needed when iron is abundant
46
what happens when iron is low vs high?
**low**: IRP and IRE bind to stabilize transferrin mRNA and increases its half life
**high**: iron blocks IRP/IRE interaction and mRNA is degraded
47
what do IRP/IRE do when they are in 5'UTR?
regulate translation in response to iron levels
48
what does ferrettin do and when would it be harmful to use?
stores excess Fe
harmful during Fe starvation by interfering with use of what little Fe is available
49
what is used to downregulate translation of proteins that require iron-based cofactors (like iron sulfur cluster enzymes like aconitase)?
IRP/IRE
50
x chromosome inactivation vs imprinting
inactivation: is random
imprinting: either one or the other is ALWAYS inactivated
51
being haploid or triploid for most chromosomes is lethal in..........
mammals
52
how can males have only 1 copy of X, while females have 2?
-in female mammals, one X is randomly selected for inactivation in embryo cells (lyonization)
-inactivated X becomes heterochromatic barr body
53
XIST
17 kb non-coding RNA located on XIC locus
NOT A PROTEIN
coats entire X-chromosome
expressed from inactivated genes via DNA methylation and nucleosome modification
binds and crawls its way down chromosome, inactivating it
54
how could you get a male calico cat?
coat color genes are on the X chromosomes
Kleinfelder= XXY
genetically male because of sex-determining genes on Y
55
women who only have 1 X chromosome are ..... but are almost always ......
viable
sterile and have other issues
56
genetics of women with 1 X chromosome
45 chromosomes
XO
57
if one X chromosome is turned off in normal females, and males manage to be fertile and normal with one copy of X, why are XO females sterile?
dont turn off ALL genes on X chromosome (on outside)
58
pseudoautosomal region
-region is similar in X and Y chromosomes and allows pairing during meiosis
- genes here are activated in normal females
59
how many copies of pseudoautosomal region in normal males, females, and Turners?
female: 2 active copies of genes in the region
male: 2 active copies of geens (one from X one from Y)
Turners (XO): 1 copy (half dose of genes --> symptoms of Turners)(2 copies are required for functional ovaries)
60
do pseudoautosomal regions reset?
yes, in germline @ meisosis
61
snRNA
gRNA
miRNA
snRNA: mRNA splicing, not encoded in genome
gRNA: RNA editing in trypanosomes
miRNA: gene expression, encoded in genome, make long transcripts, processed by Drosha
62
what is Drosha?
RNAse 3
60-70 nt long
Pri ---> pre
63
how many miRNA genes do humans have?
how many genes can 1 miRNA targe?
>1000 miRNA genes
target 400 genes
64
where are miRNA present?
in most eukaryotes (not in S. cerevisiae)
65
what cleaves pri-miRNA and pre-miRNA
Drosha cleaves pri to make pre: 60-70
Dicer cleaves pre to make final: 22 nt miRNA
66
siRNA
-processed by DICER and RISC complex
-used in biotech to lower gene expression 50-90% (make synthetic RNA to destroy)
-prevent translation
- involved in heterochromatin formation
67
expression of miRNAs using .......... enables cell specific down-regulation of target gene expression
cell specific promoters
68
Onpattro
RNAi drug for liver disease
make amyloid to block RNA
69
RITS
RNA silencing machinery recruited to heterochromatin nucleation sites
1. RITS binds to RNA transcribed from repetitive DNA
2. RDRC synthesizes complementary strand --> DS RNA
3. dicer chops ds RNA to form siRNA
4. siRNA binds to ARC and removes one strand
5. ARC binds to RITS, and siRNA directs complex to more nucleation sites and expands heterochromatin
70
lncRNA
-has transcripts from RNA pol1,2,3 RNAs from processed introns
-annotation hard because many isoforms and interleaved relationships with other genes
-associate with chromatin-modifying complexes, transcribed form enhancers, and nucleate phase separation of nuclear condensates and domains
-evolve more rapidly than protein-codings equences
-cell type specific and regulate many aspec of cell differentiation and development and other physiological processes
