X chromosome inactivation

SIGNIFICANCE: Normal females have two X chromosomes, and normal males have one X chromosome. In order to compensate for the potential problem of doubling of gene products in females, one X chromosome is randomly inactivated in each cell.

The History of X Chromosome Inactivation

In 1961, Mary Lyon hypothesized that gene products were found in equal amounts in males and females because one of the X chromosomes in females became inactivated early in development. This hypothesis became known as the Lyon hypothesis, and the process became known as Lyonization, or X chromosome inactivation. Prior to this explanation, it was recognized that females had two X chromosomes and males had only one X chromosome, yet the proteins encoded by genes on the X chromosomes were found in equal amounts in females and males because of dosage compensation.

The principles of inheritance dictate that individuals receive half of their chromosomes from their fathers and the other half from their mothers at conception. Therefore, a female possesses two different X chromosomes (one from each parent). In addition to hypothesizing the inactivation of one X chromosome in each cell, the also implies that the event occurs randomly. In any individual, approximately one-half of the paternal X chromosomes and one-half of the maternal X chromosomes are inactivated. Thus, females display a mosaic condition since half of their cells express the X chromosome genes inherited from the father and half of their cells express the X chromosome genes inherited from the mother. In fact, this situation can be seen in individuals who inherit an allele for a different form of a protein from each parent: Some cells express one parent’s protein form, while other cells express the other parent’s protein form.

Prior to Lyon’s hypothesis, it was known that a densely staining material could be seen in cells from females that was absent in cells from males. This material was termed a “Barr body,” after Murray Barr. Later, it was shown that Barr bodies were synonymous with the inactivated X chromosome. Other observations led scientists to understand that the number of Barr bodies in a cell was always one less than the number of X chromosomes in the cell. For example, one indicated the presence of two X chromosomes, and two Barr bodies indicated the presence of three X chromosomes.

Clinical Significance

The significance of Barr bodies became apparent with the observation that females lacking one Barr body or possessing more than one Barr body developed an abnormal appearance. Particularly intriguing were females with Turner syndrome. These females possess only one X chromosome per cell, a condition that is not analogous to normal females, who possess only one functional X chromosome per cell as a result of inactivation. The difference in the development of a female and a normal female lies in the fact that both X chromosomes are active in normal females during the first few days of development. After this period, inactivation occurs randomly in each cell, as hypothesized by Lyon. In cases in which inactivation is not random, individuals may have a variety of developmental problems. Therefore, there is apparently a critical need for both X chromosomes to be active in females in early development for normal development to occur.

It is equally important that there not be more than two X chromosomes present during this early development. Females possessing three X chromosomes, and therefore two Barr bodies, are sometimes called superfemales or metafemales because of a tendency to be taller than average. These females are also two to ten times more likely to suffer from mild to moderate intellectual disability.

The same phenomenon has been observed in males who possess Barr bodies. Barr bodies are not normally present in males because they have only one X chromosome. The presence of Barr bodies indicates the existence of an extra X chromosome that has become inactive. Just as in females, extra X chromosomes are also expressed in early development, and abnormal amounts of gene products result in abnormal physical characteristics and mental retardation. Males with Klinefelter syndrome have two X chromosomes and a Y chromosome. In cases in which males have more than two X chromosomes, the effects are even more remarkable.

Mechanism of X Inactivation

While it has been apparent since the 1960s that X inactivation is required for normal female development, the mechanism has been elusive. Only with the development of techniques to study the molecular events of the cell and its chromosomes has progress been made in understanding the process of inactivation. One process involved in turning off a gene (thus “shutting down” the process of transcription) is the alteration of one of the molecules of DNA known as cytosine. When a methyl group is added to the cytosine, the gene cannot produce the RNA necessary to make a protein. It is thought that this methyl group blocks the proteins that normally bind to the DNA, so that transcription cannot occur. When methyl groups are removed from cytosines, the block is removed and transcription begins. This is a common means of regulating transcription of genes. Methylation is significantly higher in the inactivated X chromosome than in the activated X chromosome. As the genes on the chromosome become inactive, the chromosome condenses into the tightly packed mass known as the Barr body. However, the process of alone cannot entirely account for inactivation.

A region on the X chromosome called the X inactivation center (XIC) is considered the control center for X inactivation. In this region is a gene called X inactive specific transcript (non-protein coding), or XIST. At the time of its discovery, this gene was the only gene known to be functional in an inactivated chromosome. It produces an RNA that remains inside the nucleus.

Evidence in humans supports the hypothesis that the XIST gene is turned on and begins to make its RNA when the egg is fertilized. Studies with mice have shown that RNA is produced, at first, in low levels and from both X chromosomes. It has been shown in mice, but not humans, that prior to inactivation, Xist (lowercased when referring to mouse genes) RNA is localized at the XIC site only, thus suggesting a potential role prior to actual inactivation of the chromosome. At this point, one X chromosome will begin to increase its production of XIST RNA; shortly thereafter, XIST RNA transcription from the other X chromosome ceases. It is not clear how XIST RNA initiates the process of inactivation and condensing of the inactive chromosome, but XIST RNA binds along the entire length of the inactive X chromosome in females. These results suggest that inactivation spreads from the XIC region toward the end of the chromosome and that XIST RNA is required to maintain an inactive state. If a mouse’s Xist gene is mutated and cannot produce its RNA, inactivation of that X chromosome is blocked. Other studies have suggested that a product from a nonsex chromosome may interact with the XIC region, causing it to remain active. As expected, but not explained, the XIST gene is repressed, or expresses XIST RNA at only very low levels, in males with only one X chromosome.

No difference has been detected between maternally and paternally expressed XIST genes in humans. This has led scientists to suspect that XIST gene RNA may not be responsible for determining which X chromosome becomes inactivated. They know it plays a role, but the extent of which remains unknown. It is also not clear how the cell knows how many X chromosomes are present. The search for other candidates for these roles is under way. Finally, there are a few genes besides the XIST gene that are also active on the inactive X chromosome. How they escape the inactivation process and why this is necessary are also questions that must be resolved.

Key terms

• Barr bodya highly condensed and inactivated X chromosome visible in female cells as a darkly staining spot in a prepared microscope slide
• dosage compensationan equalization of gene products that can occur whenever there are more or fewer genes for specific traits than normal
• mosaican individual possessing cells with more than one type of genetic constitution
• sex chromosomesthe X and Y chromosomes; females possess two X chromosomes, while males possess one X and one Y chromosome

Bibliography

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