Different unequal cross-over events between NCF1 and its pseudogenes in autosomal p47phox-deficient chronic granulomatous disease

• CGD with mutations in NCF1 is often due to unequal cross-over with NCF1 pseudogenes.
• These cross-over events can take place between or within these (pseudo)genes.
• MLPA showed 2 cross-overs within NCF1, between exons 2 and 4 and between exons 4 and 6.
• Total NCF1 deletion is most common: allele frequency in 43 CGD families was 74.4%.
• Cross-over between exons 4 and 6 in 19.2% and between exons 2 and 4 in 6.4%

Chronic granulomatous disease (CGD) is a rare congenital disorder in which phagocytes cannot generate superoxide (O2−) and other microbicidal oxidants due to mutations in one of the five components of the O2−-generating NADPH oxidase complex. The most common autosomal subtype of CGD is caused by mutations in NCF1, encoding the NADPH subunit p47phox. Usually, these mutations are the result of unequal exchange of chromatid between NCF1 and one of its two pseudogenes. We have now investigated in detail the breakpoints within or between these (pseudo) NCF1 genes in 43 families with p47phox-deficient CGD by means of multiplex ligase-dependent probe amplification (MLPA). In 24 families the patients totally lacked NCF1 sequences, indicating that in these families the cross-over points are located between NCF1 and its pseudogenes. Six other families were compound heterozygous for a total NCF1 deletion and another mutation in NCF1 on the other allele. In 8 families, the patients lacked NCF1 exons 1–4 but had retained NCF1 exons 6–10, indicating that a cross-over point is located within NCF1 between exons 4 and 6. Similarly, in 4 families a cross-over point was located within NCF1 between exons 2 and 4. Similar cross-overs, in heterozygous form, were observed in family members of the patients. Several patients were compound heterozygous for total and partial NCF1 deletions. Thus, at least three different cross-over points exist within the NCF1 gene cluster, indicating that autosomal p47phox-deficient CGD is genetically heterogeneous but can be dissected in detail by MLPA.