Lecture 27 Flashcards
where do histone modifications occur?
on N-terminal ends of core histones that protrude from nucleosome
4 examples of histone modifications
- acetylation
- methylation
- phosphorylation
- ubiquitination
3 types of histone modifiers
- Writers
- Erasers
- Readers
what are readers?
can recognize and bind modifications –> coupled with writers and erasers –> 1 mark is made which recruits a reader to allow more marks by the associated writer
how does histone acetylation increase transcription? (2)
- lysine is acetylated to neutralize its (+) charge
- loosens its interactions with (-) DNA
- chromatin relaxes, more accessibility, more transcription - makes binding site for histone code readers that promote activation
what is the writer enzyme for histone acetylation?
HAT
what proteins typically have HAT activity?
transcriptional co/activators have HAT activity
what is the eraser enzyme for histone acetylation?
HDAC
what proteins typically have HDAC activity?
transcriptional co/repressors have HDAC activity
how does histone methylation reduce transcription? (2)
- lysine or arginine is methylated (charge is unaffected) to silence genes
- makes binding site for histone code readers
what is the writer enzyme for histone methylation?
HMTase
does histone methylation promote heterochromatin or euchromatin formation?
heterochromatin
how does histone methylation promote heterochromatin formation? (4 steps)
- Heterochromatin Protein 1 (HP1) binds methylated histones (H3K9Me)
- HP1 induces methylation formation and recruits more HMTase
- HMTase methylates neighbouring nucleosomes which become new HP1 binding site
- heterochromatin spreads!
what stops the spread of heterochromatin?
- HATs bind a barrier insulator DNA seq
- this causes acetylation to counteract the spread of heterochromatin (i.e. sets limit for where heterochromatin is)
definition of epigenetics
study of heritable traits that are not explained by changes in DNA sequences due to proteins associated with DNA
can epigenetics be inherited?
yes, can be passed to daughter cells in mitosis
how do epigenetics marks get inherited in mitosis?
- during DNA replication, nucleosomes disassemble to let replication fork pass thru
- then, nucleosomes reassemble on daughter strands as a mix of new and old histones
- old histones with modifications recruit readers and writers for modifications of new histones
what is the phenomenon that occurs in Drosophila due to epigenetic modifications?
Position-Effect Variegation
what is the gene involved in position-effect variegation?
white gene –> ABC transporter that brings red pigment to eye
WT = red pigment
mutant = white pigment
what chromosome is the white gene found on?
X chromosome
describe a normal, WT drosophila eye
there is a barrier insulator, where part is heterochromatin and the other part is euchromatin
- the WT white gene is in euchromatin so it is expressed normally and the eye is fully red
describe a drosophila eye with position-effect variegation
mutagenized with X-rays –> creates inversions where white gene is close to heterochromatin
the barrier insulator has moved, so the heterochromatin can randomly spread into the white gene
–> in some cells, there will be more heterochromatin spreading so the gene will be silenced (white)
–> in other cells, there will be less heterochromatin spreading so the gene will be expressed (red)
all cells have the same genotype but different amounts of heterochromatin
what is non-mendelian genetics?
traits don’t segregate according to Mendel’s law –> genotype =/= expected phenotype
when does dosage compensation occur?
in FEMALE mammalian embryo at 8-32 cell stage
what is dosage compensation?
each cell randomly inactivates one of the X-chromosomes and forms a heterchromatic Barr body
what is the purpose of dosage compensation?
allows males and females to have equal expression of genes on the X chromosome
why do all descendent cells have the same inactive X chromosomes as the parent cell?
epigenetics are inherited in mitosis
describe dosage compensation in tortoiseshell cats
females are heterozygous with 1 allele for orange fur and 1 allele for black fur
- at 8-32 cell stage, one of the alleles is inactivated
- cat ends up as a mix of both colours
(males are all orange or all black)
what causes red-green colour blindness?
mutation in OPN1-LW gene on X chromosome
who can get red-green colour blindness?
males: mut/Y –> always
females: mut/X –> sometimes
why can some mut/X females be colourblind and others are normal?
due to X-inactivation –> can inactivate mut or WT X chromosome
describe 50-50 X-inactivation
half the cells inactivate the mutant allele, so half the cells are WT –> normal vision
describe skewed X-inactivation
more than half the cells inactivate WT so most cells are mutant –> colour blind
how does skewed X inactivation affect females?
skewed X inactivation causes female carriers of X-linked mutations to display partial symptoms
why are some X-linked diseases only found in females?
for some diseases, mut/Y males are not viable
need 1 viable gene copy to survive
what causes X inactivation in mammals
1 X is repressed with increased H3K9Me and DNA methylation, decreased acetylation
how do flies achieve dosage compensation?
male X is hyperactive
how does the male X in flies become hyperactive?
increased acetylation
what enzyme is responsible for DNA methylation? how does it work?
DNMT enzyme adds methyl to C5, mainly at CpG
what % of CpG are methylated?
60-80% in specific pattern throughout genome
where are CpGs typically located?
at intergenic regions –> correlated to heterochromatin
what are CpG islands?
CpG-rich clusters near promoters for 60% of genes
are CpG islands methylated or unmethylated? what does this indicate about the transcription activity at these sites?
unmethylated –> transcriptionally active
2 ways that DNA methylation causes gene silencing
- DIRECT EFFECT: methylation blocks TF binding
- INDIRECT EFFECT: methylation recruits HDACs and HMTs that cause repressive histone modifications
is DNA methylation inherited?
yes, it is passed on thru mitosis
describe the inheritance of DNA methylation in mitosis
- after replication, the 2 new daughter stands are HEMIMETHYLATED –> old strand has Me, new strand does not have Me
- DNMT have high affinity for hemimethylated sites so they add Me on unmethylated strands
what allows DNMT to have high affinity for hemimethylated sites?
methylation pattern on parental strand
role of Igf2 gene?
stimulates cell growth
is Igf2 sex-linked or autosomal?
autosomal
what happens to Igf+/- mice?
they are small ONLY if mutant allele was inherited from father
what is genomic imprinting aka?
silencing, Monoallelic Inheritance
what is genomic imprinting?
only 1 of the paternal or maternal copy is expressed as if there was only 1 copy of the gene in the cell
why is genomic imprinting sex-specific?
the allele that gets expressed depends ONLY on the sex of the parent it came from
what causes the imprinted allele to be silenced?
inactivated by methylation
why is genomic imprinting inheritable?
methylation is inheritable, so all daughter cells will have the same mark
what other gene is involved in Igf2 imprinting? why?
H19
they are adjacent
describe the genomic imprinting with Igf2 and H19 in the MATERNAL allele
- Imprinting Control Region (ICR) is unmethylated so CTCF binds
- CTCF acts as an enhancer-blocking insulator
- Enhancer can only reach H19 which is close by
Igf2 is maternally imprinted and only H19 is expressed
describe the genomic imprinting with Igf2 and H19 in the PATERNAL allele
- ICR is methylated so CTCF cannot bind and TF binds at enhancer
- H19 promoter is also methylated so its transcription is silenced
H19 is paternally imprinted and only Igf2 is expressed
why is the Igf2/H19 imprinting sex-specific?
the methylation at ICR is sex-specific –> only occurs in makes
when are maternal/paternal imprints established?
during gametogenesis
describe establishment of imprints in males
- original DNA methylation is removed in germ cells
- ICR is methylated
- all sperm receive the methylation
describe establishment of imprints in females
- original DNA methylation and X inactivation is removed in germ cells
- ICR is not methylated
- all oocytes are unmethylated
2 possible reasons why female ICR does not get methylated?
- missing enzyme
- gets methylated then removes the marks
why are uniparental embryos/cloned organisms INVIABLE?
researcher removes marks from established donor cell but when it develops it will only have male OR female pattern bc only 1 pattern to influence the marks –> organism is unstable
normal embryos have both patterns differentially expressed in different cells
difference btwn differentially imprinted maternal and paternal alleles
genetically identical but epigenetically different
what happens if there is a mutation in an imprinted gene?
unaffected bc gene normally silenced
what happens if there is a mutation in a non-imprinted gene?
affected bc gene normally expressed
what causes Silver-Russell Syndrome (dwarfism)?
faulty imprinting
describe the faulty imprinting in individual affected with Silver-Russell Syndrome?
paternal ICR does NOT get methylated so even paternal alleles will only expresse H19, not Igf –> i.e. have another maternal allele copy
