X-chromosome Inactivation & Dosage Compensation

Dosage Compensation

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  A phenomenon of neutralization of differences in sex-linked gene doses, i.e. males with one X-chromosome and females with two X-chromosomes have identical X-linked gene products
• First report of this phenomenon → Muller (1931) while working with the w^a allele observed that there is a mechanism compensating for the difference in the dosage of X-linked gene between the sexes
• This has been shown true for all X-linked genes with the exception of only a few alleles
• Dosage compensation in Drosophila does not operate through female X-chromosome, i.e. inactivation of one female X-chromosome that is seen in mammals in not observed in case of Drosophila
• Evidence:
• absence of phenotypic mosaicism
• Kazazzian experiment → Pgd electrophoretic variants
• Instead operates through increased transcription of the male X-chromosome
• Evidence:
• interspecific hybrid experiment of Dobhzansky (1955) D. insularis xx D. tropicalis
• The single (haploid) male X-chromosome was twice as wide as the single (haploid) femakes X-chromosome → the male X-chromosome appears puffy and diffuse, it apparently works twice as hard as the single female X-chromosome
• Transcription autoradiographic experiements of Mukherjee and Beermann (1965) suggested that the rate of transcription of the haploid male X-chromosome is 43% faster than that on a single female X-chromosome
• The puffy and diffuse appearance of the male X-chromosome is an accepted cytological marker of dosage compensation
• confirmed through measurement of steady level of transcripts of various genes
• The primary determinant of the dosage compensation pathway is the X:A ratio → changing the X:A ratio results in alteration of X-chromosome transcription
• The X:A ratio operates through Sxl gene
• Evidence:
• X-chromosome transcription is altered in Sxl mutants
• Loss-of-function in females → X-chromosome is bloated and shows increased transcription
• Gain-of-function in males → X-chromosome is condensed and transcription is reduced
• Dosage compensation is not achieved through the genes of the sex determination pathway
• Evidence:
• w^a/w^b; dsx/dsx and w^a/Y; dsx/dsx females and males are intersexual but show female and male levels of X-chromosomr transcription
• The target of Sex lethal (sxl) is a set of five known dosage compensation genes:
1. male lethal (mle)
2. male-specific lethal-1 (msl-1)
3. male-specific lethal-2 (msl-2)
4. male-specific lethal-3 (msl-3)
5. males absent on the first (mof)
• Evidence:
• male lethal phenotype
• rate of transcription, only 65% of WT male X-chromosome
• chromosome appears narrow
• genetic interdependence → mutation in any one of the msls results in male lethality, i.e. they must be functioning cooperatively in a complex → shown to be true since they co-immunoprecipitate, complex termed as "Compensasome"
• Dosage compensation in Drosophila involves a male specific histone acetylation (16Lys-H4)
• Two non-coding RNAs (products of rox-1 and rox-2 genes) are also involved in dosage compensation in Drosophila

Regulation of Dosage Compensation in Drosophila

• There are 3 essential aspects in the process:
• remodeling the chromatin of the male X-chromosome through the assembly of the multi-protein complex
• ensuring that the complex is directed to the correct target, i.e. the male X-chromosome
• ensuring sex-specific expression of this complex, i.e. the complex should be functional only in the male

Dosage Compensation Regulators and their Function

Broad Function of Gene Product	Gene	Type of Product
Transcriptional Regulation through chromatin remodeling	male-specific lethal-2 (msl-2)	RING finger protein which interacts with msl-1 and is expressed only in males
	male-specific lethal-1 (msl-1)	a novel acidic protein that interacts with msl-2 and msl-3
	male-specific lethal-3 (msl-3)	a protein with a chromodomain motif that interacts with msl-1
	maleless or male lethal (mle)	ATP-dependent helicase that interacts with rox
	males absent on the first (mof)	male-specific histone acetyltransferase
	JIL-1	a kinase which is possibly involved in nucleosomal modification
Targeting to X-chromosome	RNA on the X-1, 2 (rox-1 and rox-2)	an RNA transcript of an X-linked gene
Sex-specific expression	Sex lethal (Sxl)	an RNA-binding protein which negatively regulates msl-2