Mapping by Gene Dosage

A conceptually related approach also takes advantage of structurally rearranged chromosomes for gene mapping but does not rely on having to first segregate the abnormal chromosomes into somatic cell hybrids. This approach depends on detection of dosage differences in either gene products or the gene sequences themselves between patients, cell lines containing different numbers of copies of a particular gene. Although this method requires very careful analysis and interpretation, it has been used to assign genes to or exclude them from a region involved in a duplication or deletion. It was originally used to assign genes to chromosome 2l by detecting levels of enzyme activity in cell lines from patients with Down syndrome (three doses of chromosome 2L) that were higher than levels in cell lines from chromosomally normal Persons (two doses). At the DNA level the dosage approach has been used increasingly to assign DNA markers to the X chromosome (by comparing DNA dosage in persons with one [ie., a normal male karyotype] to five [ie., a 49, XXXXX karyotype] X chromosomes) (Fig 1) or to small regions bf a particular chromosome (by examining collections of patients with partial trisomies [three copies] or monosomies [one copy];.

One of the most direct applications of mapping by gene dosage is the-regional assignment of X-linked disease genes by examining males with cytogenetically detectable deletions of part of the X chromosome. In one well-studied case, a boy (B. B.) with no known history of any genetic disease presented with four normally distinct -X- linked conditions: Duchenne muscular dystrophy (DMD), chronic granulomatous disease (CGD), retinitis pigmentosa (RP), and a rare red blood cell phenotype. Careful cytogenetic analysis revealed a small but detectable deletion in band Xp21 (Fig 2). The coexistence of four single-gene disorders with the small chromosome deletion led to the conclusion that the genes for these four X- linked traits mapped to the deleted interval .Extension of this type of analysis to other individuals simultaneously affected with multiple X-linked diseases has allowed the regional assignment of a number of genes in this region of the X chromosome (Fig. 6). The case of B. B. turned out to be even more significant for medical genetics because (as described more fully later in this chapter) his deleted X chromosome was used directly to allow cloning of the genes for both DMD and CGD. This provides yet another example of how recognition of the unusual in medicine-in this case, the occurrence of multiple genetic diseases in a single individual - can provide important new information about normal genes, their organization, and their function.

Figure 1. Examples of mapping by dosage analysis. DNA probe 1 used at the left can be mapped to the X chromosome because the intensity of hybridization appears to be a function of the number of Xs present in each DNA sample. DNA probe 2 shows the same intensity hybridization in lanes 1 to 3, but is missing from lane 4. This locus maps to the Y chromosome.

Figure 2. Regional localization of X-linked genes in patients with X chromosome deletions. Correlation of the extent of the cytogenetic deletion with the particular disorders present in each case allows fine mapping of the individual disease genes to particular regions on the X Chromosome DMD = Duchenne muscular dystrophy, OTC : ornithine transcarbamylase deficiency, CGD : chronic granulmatous disease, RP: retinitis pigmentosa, GKD: glycerol kinase deficiency, AHC: congenital adrenal hypoplasia.