Two cases of misinterpretation of molecular results in incontinentia pigmenti, and a PCR-based method to discriminate NEMO/IKK? dene deletion

Abstract

Familial incontinentia pigmenti (IP) is a rare X-linked dominant disorder that affects ectodermal tissues. Over 90% of IP carrier females have a recurrent genomic deletion of exons 4-10 of the NEMO (IKBKG-IKKgamma) gene, which encodes a regulatory component of the IkB kinase complex, required to activate the NF-kB pathway. In IP, mutations in NEMOlead to the complete loss of NF-kB activation creating a susceptibility to cellular apoptosis in response to TNF-alpha. This condition is lethal for males during embryogenesis while females, who are mosaic as a result of X-inactivation, can survive. Recently, a second nonfunctional copy of the gene, DeltaNEMO, was identified, opposite in direction to NEMO in a 35.5-kb duplicated sequence tract. PCR-based detection of the NEMO deletion is diagnostic for IP disease. However, we present instances in which ex 4-10 DeltaNEMO pseudogene deletion occurs in unaffected parents of two females with clinically characteristic IP. These were missed by the currently standard PCR-based method, but can be easily discriminated by a new PCR-based test reported here that permits unambiguous molecular diagnosis and proper familial genetic counseling for IP.

Alterations of the IKBKG locus and diseases: an update and a report of 13 novel mutations

“…It contains exons 3-10 and carries the two MER67B repeats, which also may recombine [Aradhya et al, 2001a]. Although no evidence of an involvement of the IKBKG pseudogene in human diseases has so far been reported, the nonpathological exon 4_10 deletion of IKBKGP has been found in two families, in unaffected parents of females with a clinical diagnosis of IP [Bardaro et al, 2003]. Those cases support the need to discriminate between deletions occurring in IKBKG or in its pseudogene to perform a correct molecular diagnosis of IP.…”

“…Therefore, it will be worthwhile to undertake complete molecular analysis and so provide genetic counseling to identify carriers and to perform prenatal diagnosis. However, pre-and postnatal diagnosis of IP and EDA-ID are markedly facilitated by the combined use of PCR tests [Bardaro et al, 2003;Steffann et al, 2004]. Given the severity and familial nature of IP and EDA-ID disorders, prenatal diagnosis has particular relevance because affected IP females are at risk of having an IP-or EDA-ID-affected child.…”

“…A possible explanation for the apparent lack of mutations in the remaining 27% of IP is the detection system employed for mutational screening. Almost all investigations have been carried out by PCR-mediated strategies and subsequent analysis by automated direct sequencing [Bardaro et al, 2003;Fusco et al, 2004]. Large and complex rearrangements of the IKBKG gene (e.g., deletion, inversion, and more complex rearrangements) could escape identification by PCR-based screening.…”

Clinical Utility Gene Card for: incontinentia pigmenti

“…Long-range PCR using two specific primers able to detect the pathogenical IKBKG deletion (IKBKGexon4_10del) in the gene and able to discriminate it from the non pathogenical pseudogene deletion (PIKBKGexon4_10del). 9,10 Indeed, a non-functional copy of the IKBKG gene is located (99% identity with the gene) in the IP locus. 11 No evidence of an involvement of the PIKBKG pseudogene in human diseases has so far been reported.…”

“…12 Those cases support the need to discriminate between deletions occurring in gene or in pseudogene to perform a correct molecular diagnosis of IP. 9 A possible way to discriminate between IKBKG and PIKBKG deletions consists in testing the X inactivation pattern in white blood cells from female carriers. Conversely to PIKBKG deletions, IKBKG deletions are almost consistently associated with a full X inactivation skewing, at least after one year of age.…”