VL 9: X chromosome inactivation Flashcards
What is X inactivation and why is it needed?
Sex determination creates a need for X inactivation
Purpose: Ensures equal expression of X-linked genes in males (XY) and females (XX).
*If X creates a certain number of Proteins, female would have the double amount of proteins. But thats not the case –> X inactivation.
Mechanism: In female mammals, one copy of the X chromosome in each cell is epigenetically inactivated.
Example: Fur color in female cats showing black and orange patches due to X inactivation. Only female cats can have 4 colors.
What is dosage compensation?
Dosage compensation refers to a regulatory mechanism that ensures the equalization of X-linked gene products in males and females
Mechansims:
* Upregulation of the single X in males:
In some species, such as Drosophila, the single X chromosome in males is upregulated to match the expression levels of females’ two X chromosomes.
* X chromosome inactivation in females (Mammals): One of the two X chromosomes in females is randomly inactivated to equalize gene expression with males.
*Without dosage compensation, females would produce twice as many X-linked gene products as males, leading to an imbalance in protein production and potentially disrupting cellular and physiological functions.
What is the baseline in dosage compensation?
The baseline refers to the expected level of gene expression from X-linked genes relative to autosomal genes (genes on non-sex chromosomes) in the absence of any compensation mechanisms.
Measurement: Using Microarrays/RNA-seq provide expression of all genes in the genome to determine the X/A ratio.
- Male Baseline : X-linked expression/autosomal expression X/AA=0.5 → no compensation –> upregulatin two fold to 1
- Female Baseline: X-linked expression/autosomal expression XX/AA=1
- The latter seems to be the case in mammals
With Compensation: Adjusts male X/AA to 1, matching females.
Barr Body (1949)
Discovered by Murray Barr in 1949
Definition: Inactive X chromosome in female cells, visible as a condensed structure.
Rule:
Number of Barr bodies (#BB) = Number of X chromosomes (#X) - 1.
Lyon Hypothesis
Proposed by Mary Lyon in 196. Lyonization is the process by which one of the two X chromosomes in female mammals is randomly silenced to achieve dosage compensation between males (XY) and females (XX).
Lyon’s hypothesis explained the presence of a Barr body (condensed, inactive X chromosome) in female cells and the mosaic phenotype observed in heterozygous females.
-
random Inanctivation.
In females one of the X chromosomes is randomly selectes for inactivation in each cell. This random inactivation means that some cells will express genes from the maternal X, while others will express genes from the paternal X, leading to a mosaic pattern of gene expression. -
inactivation occurs early in development.
around the blastocyte stage -
Inactivated state is stably inherited an epigentic phenomenon.
Once an X chromosome is inactivated in a cell, it remains inactive in all descendant cells. This stable inheritance ensures that the same X chromosome remains silenced throughout the organism’s life. - Reactivation occurs in germ cells during oogenesis.
Mechanism of Lyonization
Lyonization=X-inactivation
X-Inactivation Center (Xic):
–> Master switch locus
* Silencing X chromosome in cis
* Ensuring correct and appropriate initiation of random X inactivation
* Xic produces lncRNA Xist (X-inactive-specific transcript)
* Binds in cis and accumulates along length of chromosome
The X inactivation center (Xic) is a cis-acting region on the X chromosome that is necessary and sufficient to ensure that only one X chromosome remains active
Tsix:
–> Antisense RNA to Xist, prevents Xist accumulation on the active X.
A long non-coding RNA transcribed in the antisense direction
of Xist
* Antisense transcription across Xist locus is necessary for
repression
* Initially expressed on both X chromosomes (In ES) but persists
on the active X after differentiation. It is downregulated from
the Inactive X.
* Disruption of Tsix induces X-inactivation in XY ES.
Xist RNA:
Xist is transcribed from the Xic of the chromosome that will be inactivated. Xist RNA coats the inactive X chromosome in cis (on the same chromosome from which it is transcribed), leading to its silencing.
* Xist recruits Polycomb complexes, which modify histones on the inactive X
chromosome.
* 2 x 103 Xist molecules in a female somatic cell, 6-8h half life.
* The mechanism that is responsible for
preventing Xist RNA from accumulating on the active chromosome is unknown
Note: In the lates phase, X inactivation is independet of Xist RNA
Histone Modifications:
X inactivation involves multiple levels of
chromatin modifications. The coated X chromosome undergoes various histone modifications, DNA methylation, and chromatin remodeling, resulting in a condensed, inactive state known as facultative heterochromatin.
Modification of histone tails
* Histone variants
* DNA methylation of some CpG islands
* Re-organization of higher order chromatin folding
* → stable “facultative” heterochromatin structure
Reactivation in germ cells. The inactivated X chromosome is reactivated in female germ cells during oogenesis to ensure that both X chromosomes are active in the eggs, allowing proper segregation during meiosis.
Example:
G6PD Enzyme: In heterozygous females for the G6PD enzyme gene, individual blood cells express either the G6PD+ or G6PD- allele, but not both, due to X-inactivation.
Window of opportunity
Xist RNA can establish X inactivation in
undifferentiated ES cells and during very earlycstages of differentiation, but notcsubsequently.
Mammalian X chromosome
inactivation
Female mammals:
* 1 entire X chromosome becomes
heterochromatic (visible as Barr body)
* Condensed state
* Inactive gene expression
* “facultative heterochromatin”
* N-1 rule
The N-1 rule is a principle describing the pattern of X-chromosome inactivation in female mammals. According to this rule, the number of inactive X chromosomes in a cell equals the total number of X chromosomes (N) minus one.
Abnormal Karyotypes:
* Turner Syndrome (XO):
Total X Chromosomes (N): 1
Inactive X Chromosomes: 1 - 1 = 0
Active X Chromosomes: 1
* Klinefelter Syndrome (XXY):
Total X Chromosomes (N): 3
Inactive X Chromosomes: 3 - 1 = 2
Active X Chromosomes: 1
* Triple X Syndrome (XXX):
Total X Chromosomes (N): 3
Inactive X Chromosomes: 3 - 1 = 2
Active X Chromosomes: 1
Random X inactivation: Counting
- N-1 rule
- Stochastic process: autosomal factors
promote Xist repression (blocking factors, e.g. by inducing Tsix) - X chromosomal factors activate Xist
- Blocking and activating factors compete
- checkpoint and feedback mechanisms
Imprinted X-inactivation
- skewed inactivation of the paternal X-chromosome.
- found among marsupials and in the extra-embryonic lineages of
some placental mammals. - Marsupials have an unrelated ncRNA Rsx.
- Lnx3 is a protein-coding gene from which Xist is derived (sequence
similarity).
If a normal XX female has one X inactivated, why is a X Turner female not normal?
Inactivation in normal XX female:
1. Dosage compensation
1 X chromosome is randomly inactivated to ensure dthat the dosage of X-linked gene products is equalized between males (XY) and females (XX)
2. Mosaic
random inactivation of either the maternal or paternal X chromosome resulst in a mosaic patterns of gene expression. Some express genes from maternal while others express genes from paternal.
3. Escape from X-Inactivation
Approximately 15% of genes of the inactivated X chromosome escape inactivation and are still expressed. These genes contribute to normal development and cellular function.
Turner Syndrom (XO)
occur in femles who have only 1 X chromosome isntead of 2.
- No X chromosome undergoes inactivation. The single X chromosome must compensate the absence of the second X. Leading to a shortage of X-linked gene products compared to normal XX females.
- The absence of the second X means these genes are only present in a single copy, leading to insufficient expression and contributing to the phenotypic abnormalities.
Difference:
1. Normal females benefit from the expression of certain genes from both X chromosomes, especially those that escape inactivation.Turner females lack this extra gene dosage, resulting in insufficient expression of these critical genes.
2. The absence of a second X chromosome means no potential for compensatory inactivation, leaving only one source for all X-linked genes.
This single X chromosome cannot fully compensate for the genetic content typically provided by a second X.
3. Some essential genes require dual expression (from both X chromosomes) for normal development and function.
Turner females miss out on this dual expression, leading to developmental and physiological abnormalities.
If XXY male has one X inactivated, why does he have
Klinefelter syndrome?
Klinefelter syndrome (XXY)
1. Incomplete X-Inactivation
Escape Genes: About 15% of genes on the inactivated X chromosome escape inactivation. This leads to overexpression of these genes, contributing to the KS phenotype.
2. Gene dosage effect
Extra Gene Expression: Even though one X chromosome is inactivated, the remaining active genes from the second X chromosome (including those that escape inactivation) result in a higher dosage of certain gene products compared to a typical XY male.
Impact on Development: This dosage imbalance can affect various physiological processes, leading to the characteristic features of KS.
General Effects of Extra Chromosome: The presence of an extra X chromosome (or any additional chromosome) can affect cell division, gene expression, and overall cellular function.
Interference with Normal Functions: The extra X chromosome can interfere with the normal regulation of gene expression, potentially leading to dysregulation of critical pathways.
Escape from X-inactivation
Carrel et al (2005) showed that ~15% of the genes on the inactive-X escaped from inactivation, complicating the story even further.
