lecture 24, 25 Flashcards
what does transcriptional regulation aim to regulate?
how much RNA is made
what does mRNA stability aim to regulate?
how quick RNA can degrade
what does protein stability aim to regulate?
how quick protein can degrade
what does translational regulation aim to regulate?
How much protein is made
promote-proximal elements
DNA sequences that precede the promoter of a eukaryotic gene & bind general transcription factors
enhancers
DNA sequences located far from the gene that bind transcription factors
what kinds of domains are found in eukaryotic transcription factors?
(1) DNA binding domain
(2) protein-protein interaction domain between TF-TF or TF-RNA pol
(3) domains that lead to histone modification
(4) domains that are a sensor of the physiological condition of the cell
what enhancer binds the transcription factor Gal4?
Upstream Activation Sequence (UAS)
what does UAS bind to?
the transcription factor Gal4
describe the pathway of galactose
Galactose (E) -> Galactose (I) -> Galactose-1-phosphate -> UDP galactose -> UDP-glucose -> Glucose-1-phosphate -> Glycolysis
what domains does Gal4 have?
DNA binding and activation domain
what does it mean for a transcriptional activator protein to be modular?
domains can be separated and still retain their basic activity
true or false: Gal4 is a modular activator
true
what does DNA binding domain do?
brings the activation domain to the enhancer
why is it important that the activation domain is brought to the enhancer?
so that it can help in binding to proteins associated with RNA pol
do DNA binding domains require the activation domain to bind DNA?
no; itll still bring the AD to the enhancer
does the activation domain need to have the DNA binding domain around to bind proteins associating with RNA pol?
no; itll still be able to try and bind proteins
reporter gene
a gene that is able to have its transcription state be monitored by an easily detectable phenotype associated with ON and OFF states
In cells with wild-type Gal4, what color will the colonies be in the presence of X-gal when lacZ is on?
blue
when the DNA binding domain and activation domain are separated within the Gal4 process, what happens with lacZ?
lacZ is unable to be produced and the colonies will be white in the presence of X-gal
what does Gal80 do?
binds the activation domain of Gal4
what happens when Gal80 binds with Gal4?
it prevents Gal4 from activating genes in the absence of galactose
what happens to Gal80 in the presence of galactose?
it disassociates from Gal4’s activation domain and allows it to activate genes
how does Gal80 disassociate from Gal4’s AD?
Gal3, bound to galactose, binds to Gal80, which causes it to release from Gal4
corepressor
a protein that represses transcription without directly binding to DNA
coactivator
a protein that activates transcription without directly binding to DNA
how does Gal4 help RNA pol II?
It binds basal transcription factors and the mediator complex for assistance
mediator complex
a co-activator that promotes RNA pol II activity but does not directly bind to DNA nor is it a part of RNA pol despite being required for transcription on most promoters
what types of mating type can yeast haploid cells have?
a or alpha
what type of genes do diploid cells need to suppress?
haploid, a, and alpha genes
what type of genes do a cells need to express/repress?
a and haploid genes, repress alpha genes
what type of genes do alpha cells need to express/repress?
alpha and haploid genes, repress a genes
what type of genes do diploid cells need to express/repress?
repress haploid, alpha and a genes
what proteins do diploid cells express/repress?
express a-1 and alpha-2, repress alpha-1
in a cells, what does MCM do?
acts as an activator of alpha genes
without help, what is MCM unable to do?
It cannot bind the alpha gene promoters
in alpha cells, what does MCM do?
act as an activator
are a1 proteins created in alpha cells?
no
what are the only proteins made in alpha cells?
alpha-1 and alpha-2
what does the alpha-2 protein do in alpha cells?
acts as a repressor of the MCM on the a promoter
what does the alpha-1 protein do in alpha cells?
acts as an activator on the alpha promoter
in diploid cells, what does MCM do?
acts an an activator
what type of proteins are made in diploid cells?
a1 and alpha-2
what does the alpha-2 protein do in diploid cells?
acts as a repressor and blocks MCM activity
how are haploid-specific genes and alpha-1 genes repressed in diploid cells?
by the combination of a1 and alpha-2 proteins working together
nucleosome
a protein complex; basic unit of chromatin
what are nucleosomes composed of?
8 subunits of histone (2 H2A, 2 H2B, 2 H3, 2 H4), and DNA
what are some histone variants?
CENP-A for H3 in centromeric DNA, H2A.Z to damaged DNA
histones
proteins that help package and organize DNA
what is needed for RNA pol to bind promoters securely?
the TATA box has to be free of nucleosomes
what presents a challenge for RNA pol when trying to bind promoters?
the TATA box is constantly being blocked by the nucleosome
chromatin remodeling
the process of changing histone position
what is the purpose of chromatin remodeling?
it removes histones from the promoter region which allows transcription factors and RNA pol to bind
what does H1 histone do?
stabilizes the structure of chromatin by binding to the DNA between nucleosomes, helping it pack tightly
what is acetylation
a histone modification that leads to activation of genes
what happens when lysine is acetylated?
it neutralizes the positive histone charge and reduces the interactions between histone and DNA, opening chromatin
what does HATs stand for?
histone acetyl transferases
what is HATs?
enzymes that add acetyl to histones
what does HDACs stand for?
histone deacetylases
what is HDACs?
enzymes that remove acetyl from histones
What are N-terminal tails?
the ends of histone proteins that stick out of the nucleosome
what is the purpose of post-translational histone modification?
regulates how tightly wrapped DNA is around histones which can make genes more/less accessible for transcription
what is Gal1 positively regulated by?
Gal4
what is Gal1 negatively regulated by?
Mig1-Tup1
where can Mig-1 be found?
in the nucleus when glucose is high
what is Tup1?
Tup1 is HDAC, and leads to the repression of Gal1 by creating repressive chromatin
methylation
histone post-translational modification on lysine or arginine
What does methylation of lysine do?
creates binding sites for other proteins that can activate/repress gene expression, but does not affect charge
what determines what kind of protein will bind to the methylated histone?
which histone residue is methylated and how much
even though methylation and acetylation can occur on the same amino acid, what is the restriction?
the modifications can not happen at the same time on the same amino acid
epigenetic inheritance
the passing of chromatin state from one generation to the other
what does epigenetic inheritance tell us?
during DNA replication, both the DNA sequence and the chromatin structure are passed onto the next cell generation
DNA methylation
the addition of methyl groups to DNA residues
in mammals, where is the methyl group usually added?
It is added to the cytosine in a CG dinucleotide
hemimethlyated DNA
DNA molecules methylated on only one strand
is DNA methylation or histone modifications more stable?
DNA methylation
where would you find DNA strands that have undergone methylation
in regions of the genome that are maintained in an inactive state for an organism’s entire life
synergistic effect
the effect of a combination of genes is greater than addicting the individual effects
enhanceosomes
a large protein complex and the enhancer it binds that work together to activate transcription
how are enhanceosomes formed?
by the binding of multiple regulatory proteins to the multiple binding sites in an enhancer
describe the process of enhanceosomes recruiting chromatin remodelers
(1) GCN5 binds the enhanceosome and acetylates the surrounding histone
(2) acetylated histones recruit proteins that move away histones
(3) histones move form the promoter and expose the promoter that can now bind RNA pol
what is GCN5?
a histone acetylate
insulator
a cis-acting element (DNA sequence) that limits chromatin state to a distinct DNA region
enhancer blocking insulator
DNA element that blocks the action of an enhancer
what does the action of an insulator involve?
DNA binding proteins that form complexes that can fold DNA and promote long range DNA interactions
hemetochromatin
condensed chromatin that is not favorable for transcription
euchromatin
open chromatin that is favorable for transcription
constitutive heterochromatin
permanently compact and inactive chromatin
where can you find constitutive heterochromatin?
in centromere and telomere regions
what can heterochromatin do?
change its position and spread to neighboring chromosomal regions
position effect variegation
expression of a gene depending on its position and silenced in some cells and active in others, despite being a part of the same issue as the other cells
when is position effect variegation most prevalent?
when a gene is placed in proximity to heterochromatin and caused by the spreading of repressive chromatin
what is HP1
a protein that is required for the repressed chromatin state and binds H3K9 methylated histones
what can some gene products do that effect the spread of heterochromatin?
it can bolster the spread and silence more of the gene in cells or suppress the spread and allow for more expression of previously silenced genes
barrier insulators
DNA element that blocks the spreading of heterochromatin
how do barrier insulators and HAT work together?
HAT adds another measure for euchromatin near the heterochromatin to be not as tightly packed to mitigate heterochromatin’s effect
genomic imprinting
genomic region that are repressed depending on whether they were inherited from the mother or father
paternal imprinting
repression of an allele inherited from the male parent
maternal imprinting
repression of an allele inherited from the female parent
what does genomic imprinting require to work?
insulators
what genes require enhancer function for transcription?
H19 and lgf2
what is the limitation with the enhancer with regards to H19 and lgf2?
it can activate either one, but not both at the same time
what is CTCF
a DNA-binding protein that regulates chromatin structure and gene expression
what does CTCF do?
binds the imprinting control region sequence when the DNA is unmethylated
what happens when ICR (imprinting control region) is methylated?
CTCF does not bind the DNA at ICR and activates lgf2
what happens when CTCF binds the DNA at ICR?
lgf2 is blocked and H19 is activated
which allele is methylated?
the paternal allele
which allele is unmethylated?
the maternal allele
if an allele is methylated, which gene is expressed/repressed?
lgf2 is expressed, H19 is repressed
if an allele is unmethylated, which gene is expressed/repressed?
H19 is expressed, lgf2 is repressed
what is H19
a non-coding RNA gene expressed by the maternal gene, and plays a role in regulating growth suppression
what is lgf2
a protein-coding gene expressed by the paternal gene, and plays a role in regulating growth during development
if a recessive allele is inherited from a female and is imprinted in the female, what will the phenotype of the progeny be?
dominant
if a recessive allele is inherited from the female and it is imprinted in the male, what will the phenotype of the progeny be?
recessive
if a recessive allele is inherited from the male and it is imprinted in the female, what will the phenotype of the progeny be?
recessive
if a recessive allele is inherited from the male and it is imprinted in the male, what will the phenotype of the progeny be?
dominant
x-chromosome inactivation
one copy of the X becomes transcriptionally inactive
where is Xist transcribed from?
X-inactivation center
what will Xist do?
it coats the chromosomes it transcribed from and induces a silent chromatin state
HMTase
enzyme responsible for the methylation of histones
what does H2A do?
helps to package the DNA into a structure that is wrapped around histone proteins
what does H2B do?
helps in the structural integrity of the nucleosome
what does H3 do?
regulate gene expression
what does H4 do?
regulate gene expression
