Mapping of a de novo unequal crossover causing a deletion of the steroid 21-hydroxylase (CYP21A2) gene and a non-functional hybrid tenascin-X (TNXB) gene.

In the human genome, the major histocompatibility complex class III region on chromosome 6p21.3 stands out as an area of remarkably high gene density. 2 Within this region, a section of particular complexity centres around the C4 genes, which encode the fourth component of complement. Centromeric to C4 lies the CYP21A2 gene, which encodes steroid 21-hydroxylase, a key enzyme in the biosynthesis of cortisol and aldosterone. 8 9 The TNXB gene, which encodes the extracellular matrix protein tenascin-X, lies centromeric to CYP21A2 and is transcribed from the opposite strand. Telomeric to C4 lies the RP1 gene, encoding a putative serine/threonine kinase. A typical chromosome 6 carries a duplication of an area of approximately 30 kb encompassing the entire C4 and CYP21 genes 8 plus small truncated sections of RP and TNX. This tandem repeat has been named the RCCX module after its four constituent genes. 14 16 17 In most white populations, about 70% of all haplotypes have a bimodular arrangement similar to the one shown in fig 1. The complex genetics of this region, and the activities and clinical significance of the proteins encoded here, have been the subject of several recent reviews. 18–21 Each chromosome 6 has at least one RCCX module, most have two as described above, some have three, and in rare cases as many as four contiguous RCCX modules have been found on a single chromosome. As bimodularity is the standard, publications often refer to haplotypes with one module as deletions, and to haplotypes with three modules as duplications, especially when focusing on the C4 or CYP21 genes. The overall layout of the RCCX region can be determined by short range and long range restriction mapping. Many studies have used TaqI and BglII restriction analysis of genomic DNA and comparison of the relative intensities of the bands obtained by hybridisation to C4, CYP21A2, and TNXB probes 8 22–24 to establish haplotypes in families of patients with congenital adrenal hyperplasia and in controls. This approach is usually sufficient, but in complicated cases, rare cutters such as SacII or BssHII are needed to determine the size of the entire region and, hence, the number of RCCX modules. 26 The tandem repeat structure of the RCCX region promotes the chance of misalignment during meiosis. If a crossover then occurs, it effectively removes one of the modules. Between standard bimodular chromosomes, this process joins a part of the telomeric module to its homologous counterpart in the centromeric module (see fig 5 in the discussion for a typical example). The site of such a crossover determines whether or not the remaining monomodular chromosome carries a genetic disorder. An unequal crossover between C4A and C4B is relatively harmless, because both genes express a functional C4 protein, and so does their fusion gene. Such monomodular haplotypes lacking one of the C4 genes and the CYP21A1P gene occur at a frequency of 5%–20% in the general population. 27–29 An unequal crossover between CYP21A1P and CYP21A2, on the other hand, usually generates a fusion gene that is CYP21A1P-like in its 5′ section and contains several mutations rendering it inactive (a haplotype often referred to as a CYP21A2 deletion). Absence of a functional CYP21A2 gene is one of the defects that contributes to steroid 21-hydroxylase deficiency, the cause of over 90% of all cases of congenital adrenal hyperplasia. This is a disorder of adrenocortical steroid biosynthesis which in severe cases causes life threatening salt losing crises in untreated paediatric patients. An unequal crossover between TNXA and TNXB not only eliminates the CYP21A2 gene, but may also create a non-functional TNXB/TNXA hybrid that contains a 120 bp deletion on an exon-intron boundary normally present in TNXA only, and is therefore unable to express the tenascin-X protein. This defect contributes to the Ehlers-Danlos syndrome, a recessively inherited disease of connective tissue. 30 31 The mechanisms of these crossovers are difficult to understand, because usually only the recombinational end product is available for analysis. The concept of a deletion of the CYP21A1P pseudogene as a premutation has been proposed in a report on a de novo deletion of CYP21A2 by recombination between a standard bimodular chromosome and a monomodular chromosome. More recently published studies also provide evidence that TNXB/TNXA hybrids are the result of a crossover between a bimodular and a monomodular chromosome. 17