Female Bias in Rhox6 and 9 Regulation by the Histone Demethylase KDM6A
Figure 5
Rhox6 and 9 expression and KDM6A occupancy are high in ovary where the genes are imprinted.
(A) Rhox6 and 9 have significantly higher expression in mouse ovary than in testis, based on re-analyses of published expression array data for 14 testis and 12 ovary specimens (*p<0.05, **p<0.001). Expression normalized to array mean (see also Figure 1C). (B) KDM6A occupancy measured by ChIP-qPCR at the 5′end of Rhox6 and 9 is higher in ovary than in testis, and is very low to undetectable in brain where these genes are not expressed [19]. Occupancy levels were normalized to input fractions. (C) Kdm6a has high expression in female tissues especially ovary based on analyses of published expression array data (***p<0.0001) (see also Figure 1F). (D) Rhox6 and 9 are expressed from the maternal allele only in ovary because of imprinting. DNA sequence chromatograms of gDNA and RT-PCR (cDNA) products derived from ovary from female F1 mice obtained by mating M. spretus males with C57BL/6J females with or without an Xist mutation (XistΔ and XistΔ−). SNPs to distinguish Rhox6 and 9 alleles on the active X (Xa) and on inactive X (Xi) are indicated below. In ovary from both XistΔ and XistΔ− mice the gDNA shows heterozygosity at the SNPs while the cDNA shows only the maternal allele, consistent with paternal imprinting. (E) By qRT-PCR Rhox6 and 9 are more highly expressed in ovary from XistΔ mice in which the maternal X chromosome is expressed in all cells, compared to XistΔ− mice in which there is random X inactivation (1.7-fold and 3-fold, respectively), suggesting that Rhox6 and 9 are silenced by X inactivation. Values represent the expression ratio between XistΔ and XistΔ− ovaries.