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Previous studies have shown that the chloride channel gene Clc4 is X-linked and subject to X inactivation in Mus spretus, but that the same gene is autosomal in laboratory strains of mice. This exception to the conservation of linkage of the X chromosome in one of two interfertile mouse species was exploited to compare expression of Clc4 from the X chromosome to that from the autosome. Clc4 was found to be highly expressed in brain tissues of both mouse species. Quantitative analyses of species-specific expression of Clc4 in brain tissues from mice resulting from M. spretus ؋ laboratory strain crosses, demonstrate that each autosomal locus has half the level of Clc4 expression as compared with the single active X-linked locus. In contrast expression of another chloride channel gene, Clc3, which is autosomal in both mouse species is equal between alleles in F 1 animals. There is no evidence of imprinting of the Clc4 autosomal locus. The recent finding that two closely related mouse species, Mus spretus and laboratory strains (a mixture of M. musculus musculus and M. musculus domesticus) differ in the chromosomal location of the chloride channel gene, Clc4 (1, 2) presents a unique opportunity for studying the function of the Clc4 gene product and for investigating the relationships of dosage compensation, X chromosome inactivation, and the evolution of the mammalian X chromosome. Ohno's hypothesis (3) predicted that the mammalian X chromosome as a linkage group would be preserved, Clc4 being one of the few exceptions (4). Ohno also hypothesized that during the evolutionary transition to dosage compensation X-linked genes would be up-regulated on the single active X chromosome of males and females to maintain balance with autosomal genes that are expressed from both alleles (3).Clc4 is a member of the class of voltage-gated ion channels that all share a predicted 12-transmembrane domain structure based on hydrophobicity analysis (5, 6). As expected from the DNA sequence diversity of the chloride channel encoding genes, different family members now have been shown to differ widely in terms of tissue-specific expression and association with disease phenotypes. For example, CLC1 mutations are associated with autosomal dominant or recessive myotonia (7,8); whereas, mutations of CLC5, a putative renal chloride channel gene, have been found in different kidney disorders (9). To determine the repertoire of tissue expression for the mouse Clc4 gene, Northern and reverse transcriptase-PCR (RT-PCR) analyses of mRNA were carried out in both M. spretus and laboratory strain C57BL͞6Ros. In situ hybridization was done to determine the location of Clc4 expression in embryos and adult mice.To test directly the possibility of up-regulation of X-linked transcription as predicted by Ohno, we generated backcross mice resulting from matings between the interfertile species, M. spretus and a laboratory strain C57BL͞6Ros. Expression of Clc4 from the X-linked and from the autosomal loci was measured within individual backcross ani...

A hybrid congenic strain, C57BL/6J.SPRET-Hprt(a), carrying 17 map units of Chromosome (Chr) X from Mus spretus on a background of C57BL/6J. has the novel phenotype of low fertility associated with small testis weight. In histological cross-section, many of the tubules in the testes of these congenic mice are empty except for Sertoli cells, while the other tubules appear to be normal. The gene, interspecific hybrid testis weight 1 (Ihtw1) causing this phenotype, has been fine mapped by using the strategy of generating subcongenic strains from recombinants within the congenic region. Genetic and phenotypic analysis of the subcongenic strains has defined a critical region of 1.8 map units for Ihtw1. This region of the genetic map is orthologous to the region on human Chr X containing the gene for the Borjeson-Forssman-Lehman syndrome, an inherited disease in which males show microorchidism.