Chapter 21 - Epigenetics

Question 1

What is epigenetics?

Answer 1

Change in gene expression or phenotype that are potentially heritable but that do not alter the underlying DNA base sequences

Typically, very stable from generation to generation, but can be induced by environmental factors

Question 2

What are the 3 mechanisms leading to epigenetic changes?

Answer 2

DNA methylation

Histone modification

RNA molecules

Question 3

What is DNA methylation?

Answer 3

Always on cytosine in eukaryotes - 5-methylcytosine

Associated with repression of transcription
- Methyl groups typically sit in major grooves of DNA where DNA binding proteins sit
- When methylation occurs, it becomes difficult for these proteins to bind in and control regulation
- Can also block transcription factors and polymerases from binding

Typically adjacent to guanines - forms CPG dinucleotides

Regions with many CPG nucleotides - CPG islands
- Typically located on or near promoters (affect gene regulation)

Can repress or activate genes by methylating or demethylating cytosines in genes

Question 4

What is methyltransferase and how does it work?

Answer 4

Enzyme that methylates DNA

Come in and recognize hemi-methylated DNA (one strand is methylated, other isn’t)

Will add methyl groups to unmethylated cytosines

In this way, methylation is maintained across cell divisions and future generations

Question 5

What is the royal jelly example with DNA methylation?

Answer 5

Royal jelly silences the Dnmt3 gene, which regulates DNA methylation

Typically, the gene is active in bees, which methylates their DNA, resulting in a worker phenotype

In royal jelly, the gene is shut down, which causes no methylation of the DNA, resulting in a queen phenotype

Question 6

How is DNA methylation traditionally detected?

Answer 6

Some restriction endonucleases are sensitive to methylation and will not cut a sequence that contains a methylated cytosine, while other restriction enzymes are insensitive

Can use a combo of sensitive and insensitive to cut up the DNA and then run it on a gel to see the patterns of unmethylated and methylated

Question 7

How is bisulfate sequencing used to detect DNA methylation?

Answer 7

Can treat unmethylated cytosines with sodium bisulfate, which converts them to uracil

Can sequence pretreated and treated to see where unmethylated cytosines are in DNA sequences

Cytosines that come up as thymine indicate that they were unmethylated

Cytosines that are unchanged indicate methylation

Question 8

What are histone modifications?

Answer 8

Typically occur on the tails of histone proteins

Include additions of phosphates, methyl groups, acetyl groups, and ubiquitin

Can occur at different amino acids on different histones leading to 100+ unique potential changes to the histone itself

Can either increase or decrease rate of transcription of a gene
- Do this by altering chromatin structure – more open or closed structure
- Open – increase rate of transcription
- Closed – decrease rate of transcription

Question 9

What is the effect of histone acetylation on gene expression?

Answer 9

Increases gene expression

Addition of acetyl groups on histones opens chromatin structure

Question 10

What is the effect of histone methylation on gene expression?

Answer 10

Decreases or increases gene expression

Depends on amino acid it binds to - can open or close structure

Question 11

What are polycomb groups and what do they do?

Answer 11

Large group of proteins that represses transcription by modifying histones

Often responsible for adding or removing modifications

Bind in to where histones are and make it more difficult for the enzymes/proteins that are necessary for transcription to bind into DNA

Proteins are recruited to specific targets on the DNA by transcription factors, chromatin markers, or other noncoding components

Question 12

What are the 2 hypotheses for how histone modifications are passed down through generations?

Answer 12

When old histones are distributed after replication and new histones are added, epigenetic marks in old histones recruit enzymes to make similar changes in new histones

Epigenetic marks are all lost during replication but modifying enzymes remain attached to histones and reestablish marks on both old and new histones after replication

Question 13

How are histone modifications detected?

Answer 13

Chromatin immunoprecipitation (ChIP)

Fragment the chromatin and apply a specific antibody that binds to histone probes
- Probes cause modification to chromatin itself and histones are removed
- DNA is precipitated out, and you can determine genetic location of where these histone modifications were

Long and drawn out process – now use ChIP-seq
- Can sequence fragments that are marked, so you can determine locations where modifications are occurring

Question 14

What are the 5 mechanisms in which RNA molecules lead to epigenetic changes?

Answer 14

X inactivation
- Xist - encodes a long noncoding RNA that suppresses transcription through histone methylation

Paramutation
- siRNA epigenetically alter allele that induces a change in another allele

siRNAs silence genes and transposable elements by directing DNA methylation

Methylation and histone modifications influence the expression of microRNAs

MicroRNAs control the expression of genes that produce epigenetic effects

Question 15

What are the 5 epigenetic effects and developments?

Answer 15

Cell differentiation

Paramutation

X-inactivation

Genomic imprinting

Monozygotic twins

Question 16

What is cell differentiation?

Answer 16

All cells are genetically identical in an organism, yet these cells exhibit a vast array of phenotypes
- Differences between cells are very stable and are passed on from generation to generation (cellular level) even though DNA sequence is the same

As stem cells divide and give rise to different specialized cells, the gene expression programming of the cell becomes progressively fixed
- Each cell is only expressing genes that are necessary to be an epithelial cell

Changes in DNA methylation and chromatin structure that play a role in both silencing and activation of genes that are necessary for cell-specific expression

Question 17

What is paramutation?

Answer 17

Type of transgenerational epigenetic effect in which one allele of a genotype alters the expression of another allele
- This alteration persists for several generations
- Persistence can occur even after allele is taken out of gene pool

No change in DNA sequence itself

Altered allele is able to convert another allele to new phenotype

Question 18

How is mutation seen in corn at the b1 locus?

Answer 18

Helps determine amount of purple pigment that is produced by the corn

B’ is altering allele – alters B-I allele so it is regulated/silenced
- Both alleles at DNA sequence level are identical – called epialleles

Heterozygote plant will produce light pigmentation, but it will convert B-I allele to B’*

B’* will be passed on to future generations and will be able to convert other alleles to the B’* allele just as the B’ did

Question 19

How does paramutation work at the molecular level for corn?

Answer 19

B-I locus has 7 tandem repeats upstream of coding sequence that don’t encode a protein but could act like an enhancer for transcription

Both B-I and B’ alleles have tandem repeats upstream, but chromatin structure between them is different

At B-I, chromatin structure is more open at tandem repeats/coding region
- Most likely allowing transcription to occur more regularly

At B’, chromatin structure is more closed, causing transcription to be reduced

Most likely occurs because of an siRNA that binds near tandem repeats to cause chromatin structure to close up more

Question 20

What is X-inactivation?

Answer 20

Random inactivation of one of the X chromosomes in females to provide equal expression of genes in males and females – form of dosage compensation

Once a particular X chromosome is inactivated, it will remain inactivated when DNA is replicated, and epigenetic mark on inactivated X is passed on to future generations of cells

Controlled by a segment of X chromosome called X-inactivation center

Key player is the Xist protein – encodes for long noncoding RNA
- Long noncoding RNA is transcribed by one of the X chromosomes, and it coats the X chromosome in itself
- Recruits the polycomb recessive complex I, which produces epigenetic marks (typically histone modifications) along X chromosomes that will become inactive
- Marks change configuration of chromatin structure on chromosome to repress transcription of all of the genes on the X chromosome, thereby rendering it inactive

Question 21

How does the other X chromosome not become inactive in X-inactivation?

Answer 21

Other genes at X-inactivation center play a role in not allowing the other chromosome to also be inactivated

Tsix is an antisense (opposite direction to Xist) long noncoding RNA to Xist
- Represses Xist on the X chromosome that is going to stay active

Jpx stimulates expression of Xist on the inactive X chromosome

Xite is another long noncoding RNA that stimulates the expression of Tsix on the active chromosome

Question 22

What is genomic imprinting?

Answer 22

Sex of the parent that transmits a certain allele determines the expression of an allele in its progeny

Thought to be due to methylation of alleles in the parents

Imprinted genes in mammals typically affect early embryonic development and fetal growth - Leads to genetic conflict hypothesis

Question 23

What is the genetic conflict hypothesis?

Answer 23

Suggests that there are different and conflicting evolutionary pressures acting on maternal and paternal alleles that affect fetal growth

There are paternal alleles that tend to maximize the size of the offspring, and these alleles are favored evolutionarily – associated with infant survival and adult health

Conversely, maternal alleles tend to limit/reduce fetal growth to allow for less strain on the mother to carry the progeny and allow her to reproduce again

Predicts that genomic imprinting evolves in this way

Question 24

What occurs with monozygotic twins and epigenetic effects?

Answer 24

Because monozygotic twins develop from the same egg, they’re genetically identical, however we often see variation between identical twins in terms of appearance, health, behavior, etc.

Variation isn’t well understood, but it is believed to come from epigenetics
- Could be environmental epigenetic factors or be present in one twin but not the other or can change in individual over time

Question 25

What is behavioral epigenetics?

Answer 25

Life experiences can have long-lasting effects on behavior and be passed on to future generations

Maternal behaviors in rats who lick/groom their offspring more tend to see less fearful offspring – due to DNA methylation or histone modification in future generations

Maternal behaviors in rats and children who experience abuse see an increase in epigenetic marks (DNA methylation) that affect their behavior downstream

Question 26

What are the epigenetic effects on metabolism?

Answer 26

Diets of parents can have downstream effects on their offspring

Studies have shown that there are differences in epigenetic marks based on the diet of the parent that altered metabolism and health in the offspring

Question 27

What is the epigenome?

Answer 27

The epigenome is the overall pattern of chromatin modifications in the genome

Characterizations of different cell types has revealed amazing variation in epigenetic marks

Studied through the use of epigenome-wide association studies (EWAS) and epigenetic editing
- Difference between GWAS and EWAS in terms of capabilities is that EWAS is far more limited because the genome stays constant and the epigenome constantly changes
- Can do epigenetic editing – changing chromatin structure to influence gene expression
- CRISPR/Cas9 system can make both genetic and epigenetic changes