Genomic imprinting and X-chromosome inactivation

Two examples of biological processes regulated by epigenetics:
(1) Genomic imprinting
(2) X-chromosome ____________

This is monoallelic expression of a gene in a parent-of-origin specific manner

We inherit 2 copies of each gene: _ from Mom, _ from Dad
- In most cases, both copies are active and contribute to the phenotype

_________ genes are those that are only expressed from one parental allele
- This whole process is regulated by epigenetics!

________ is the reason that we need both the biological male and biological female for successful reproduction

Nuclear Transfer Experiments (1980s)
1. Control: take sperm, take egg, fuze them, and transplant them into a mouse, which produced a normal embryo with a normal placenta

2. Take pronuclei from two EGGS, fuze them, you get a _________ embryo and a __________ placenta (lethal)

3. 2. Take pronuclei from two SPERM fuze them, you get a _________ embryo and a __________ placenta (lethal)

This experiment demonstrated that the 2 parental gametes _________ (are/are not) equivalent because of imprinting.
- __________ the dose of parental genes results in lethality (maternal or parental)

Naturally Occuring Double Dose Situations in Humans

- Parthenogenetic Conceptuses: two ________ fuze together into a Ovarian Teratoma (dermoid _________)

- An ovarian teratoma is a tumor with precursors such as teeth and bones (not a functional/viable fetus)

Differential Marking of the Parental Genomes

- There are methylated genes in sperm and egg that stay methylated throughout your entire _____ (through zygote, cell differentiation, cell division, etc.)
- At the moment of __________, both the paternal and maternal genome are demethylated -> Why?
- Embryonic stem cells in the blastocyst need to _____________, so that means they need to lay down new DNA methylation _________ that reflect the cell type they turn out to be, not possess the same methylation profile as the gametes

This is a region that is responsible for controlling allele-specific expression of imprinted genes

- They can regulate multiple imprinted genes
- Imprinted genes are typically clustered in the genome and share regulatory elements (ex: chromosome 11 has 6 imprinted genes that are regulated by one ___)

ICR: H19/IGF2

H19 is __________ expressed and encodes a ___RNA
IGF2 is __________ expressed
There is an ICR between the two genes that is ________ on the paternal allele, not the maternal allele
- Because of this, an insulator protein, ____, binds the ICR on the maternal allele, blocking the promotion of ____ via enhancers

Some ICRs control non-coding RNA expression (KCNQ1 locus)

- The ICR allows lncRNA to be made, accumulates, complexes with other proteins, and blocks __________ of paternal/maternal imprinted genes.

Imprinting Disorders On the Same Chromosomal Locus
(1) ___________ Syndrome = the disrupted allele is inherited from _________(mom/dad)
(2) __________ -______ Syndrome = the disrupted allele is inherited from _________(mom/dad)

Molecular Basis of AS and PWS

- For 70% (most cases) of AS or PWS there is a _______ on chromosome 15 from either Mom or Dad
- This means that there must be some genes that are very important for imprinting in this region of the chromosome (you may get one dose of the gene from one parent, but not the other)

- For a small number of cases, Uniparental Disomy (UPD) is the cause
- This occurs when a child recieves both copies of a chromosome from _____ parent and no copies from the other (double dose)

- __________ are linked to a small % of cases. This means that the child inherits the correct chromosomes, but one chromosome copy may be methylated to give a profile that doubles the dosage of imprinted genes (double dose)

This happens during early development, in XX females, one of the X chromosomes is inactivated (Barr ________)
- This process is RANDOM, one of the two copies of X is randomly inactivated, meaning some somatic cells may have a red X active and some somatic cells may have a black X active (example) which gives rise to ______________.

X-chromosome inactivation occurs in normal somatic cells (NOT GAMETES) of all XX females to balance gene dosage with ________, which gives rise to mosaicism.

The Lyon Hypothesis
- Mary Lyon (1925-2014)
- Studied X-chromosome Inactivation

- She proposed that X-inactivation had to happen in early __________, had to be a _________ process, and had to be heritable through ____ division

The First Cloned Cat
- Carbon Copy was cloned from a calico named Rainbow
- Despite being clones and having identical ___, their coat patterns were totally different due to _-_______

The X inactivation Center: Useful Abbreviations

1. X chromosome Inactivation = (XCI)

2. X inactivation center = (XIC)

3. Inactive X = __

4. Active X = __

Xist long non-coding RNA (Xist lncRNA)

- How does X-inactivation happen? --> It all comes down to this lncRNA called ____________.

- This gene is located within the X-_____________ center (XIC), and is expressed by the ________ _ (Xi)

How does X-inactivation Occur?

1. Xist is transcribed, but not __________ (remember its a lncRNA) on the Xi

2. Xist then _________ the Xi

3. Methylation is REMOVED due to the loss of Histone ___s

4. Xist recruits ___2 complex, causing a GAIN of Histone PTMs and switching of Histone _________.

5. The Barr Body now gains methylation at all of its promoters

Can We Correct Aneuploidy?

- As we've learned more about how Xist initiates the process of X-__________ people started to ask if we could use this as a potential THERAPEUTIC option
- Experiment: If you were to move Xist, the gene, to a different location on the X-chromosome, what would happen?
- Result: X-inactivation still happens to some extent, but you have new regions of initiation correlating with those that exist physically in close proximity

What if we used CRISPR to insert a copy of Xist into of the copies of chromosome 21 (Trisomy 21/Down Syndrome)?
- Could we silence the extra chromosome?
- Scientists created a chromosome 21 ____ Body
- Cells with the silenced extra copy began to behave more like unaffected cells!