Epigenetic regulation Flashcards
Where does methylation occur?
Methylation usually occurs at the CPG islands of the 5’ end of a gene. Usually called the proximal promotor vs the core promoter. CPG islands are present for all house keeping genes and half of tissue specific genes. Methylation usually inactivates. CPG islands are cytosine rich so those are often methylated.
Describe methylation levels and enzymes involved. Discuss the effect of methylation and writer vs reader.
So after fertilization there is a wave of demethylation. Male is quickly demethylated (demethylase dMTase) and females are slowly demethylated through subsequent divisions. Then there is denovo methylation via DNA methyltransferase which occurs during gastrulation but rarely occurs after. DNMT. These enzymes are writers. The transcription factors which come to methylated regions and either inhibit or activate are called readers.
Imprinted genes is just a constant level because those methylation patterns have already been established and do not change.
Describe Rhett syndrome.
Neurological symptoms. It is caused by a defect in MECP2 protein which is a transcription factor which inactivates genes that are methylated with particularly high activity in the brain. The thought is that overexpression of genes in the brain leads to developmental issues. It is an X linked dominant and is the most common cause of mental retardation in females. It is an autism spectrum disorder and monogenic.
What is the trend of imprinted gene methylation in the life of an organism.
It stays the same. However when sperm is made, all methylation is reset to adopt the male pattern of methylation (even maternal genes) / its not about the genes themselves but the overlying regulation (epigenetics)
For oogenesis, the methylation is of the mom.
Prader Willi Syndrome &
Angelman Syndrome
Prader Willi Syndrome: a deletion on chromosome 15 of the SNRPN and UBE3A. Since the paternal SNRPN is the only active copy in the organism, they have a lack of SNRPN protein. This leads to mental retardation, hyperphagia (insatiable hunger so theyre really fat)
Angelman syndrome: THe maternal UBE3A is the only active copy so deletion in the maternal allele leads to a lack of UBE3A protein. And this leads to mental retardation, excessive laughter, seizures.
What are post translational modifications to the histone protein that can occur to regulate transcription of the associated DNA?
acetylation: neutralizes the positive charge of the lysine on the N-terminal tail of core histones so the interaction with negative DNA is weakened.
Methylation - tightens histone binding
Two others are sumoylation and phosphorylation of serine residues.
What is noncoding RNA and what is the impact in X inactivation?
Non coding RNA are transcripts that are not made into protein which regulate transcription, chromatin remodeling, etc.
Now both chromosomes have an Xic (X inactivation centeR) With low levels of Xist transcript being made. At some point one of the X’s transcribe Tsix which is antisense to the Xist transcript. It binds and that X is the only one activated. Xist is a noncoding RNA that shuts off transcription by modifying the histones.
Whats the deal with the agouti gene?
The agouti gene and the consequent phenotypic differences is due to methylation of the CPG islands. Rats with methylation will develop normally. THose lacking methylation in the agouti gene will become fat and yellow despite being genetically identical. Diet a folic acid and stuff during pregnancy gave normal offspring so A LACK of methyl groups are a concern.
Describe the two methods used to study epigenome
Bisulfite conversion. A strand is denatured and then exposed to bisulfite. Bisulfite changes all C’s except for methylated ones to uracil. Then you reverse transcriptase (dunno why, to get rid of the U’s) and analyze the sequences. Where there is a C is where the methylation was.
Another is restriction enzyme (southern blot hybridization) You use a isochizomer (cuts the same sequence) however one is methyl insensitive (cuts right through) another is Methyl sensitive will not cut at sites where there is methylation. Then you can use a southern to look at the fragments.
