Second-site mutation in the Wiskott-Aldrich syndrome (WAS) protein gene causes somatic mosaicism in two WAS siblings

Genetics and Molecular Biology Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4442, USA.
Journal of Clinical Investigation (Impact Factor: 13.22). 06/2003; 111(9):1389-97. DOI: 10.1172/JCI15485
Source: PubMed


Revertant mosaicism due to true back mutations or second-site mutations has been identified in several inherited disorders. The occurrence of revertants is considered rare, and the underlying genetic mechanisms remain mostly unknown. Here we describe somatic mosaicism in two brothers affected with Wiskott-Aldrich syndrome (WAS). The original mutation causing disease in this family is a single base insertion (1305insG) in the WAS protein (WASP) gene, which results in frameshift and abrogates protein expression. Both patients, however, showed expression of WASP in a fraction of their T cells that were demonstrated to carry a second-site mutation causing the deletion of 19 nucleotides from nucleotide 1299 to 1316. This deletion abrogated the effects of the original mutation and restored the WASP reading frame. In vitro expression studies indicated that mutant protein encoded by the second-site mutation was expressed and functional, since it was able to bind to cellular partners and mediate T cell receptor/CD3 downregulation. These observations were consistent with evidence of in vivo selective advantage of WASP-expressing lymphocytes. Molecular analysis revealed that the sequence surrounding the deletion contained two 4-bp direct repeats and that a hairpin structure could be formed by five GC pairs within the deleted fragment. These findings strongly suggest that slipped mispairing was the cause of this second-site mutation and that selective accumulation of WASP-expressing T lymphocytes led to revertant mosaicism in these patients.

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    • "dysplasia with immunodeficiency (XL-EDA-ID; Nishikomori et al., 2004), Wiskott-Aldrich syndrome (WAS; Ariga et al., 2001; Wada et al., 2003; Stewart et al., 2007; Trifari et al., 2010), and lymphocyte adhesion deficiency-1 (LAD-1; Tone et al., 2007; Uzel et al., 2008). Although somatic reversion is infrequent overall, it has been observed in 11%, 18%, and 35% of patients with WAS (Stewart et al., 2007), Fanconi anemia (Kalb et al., 2007), and epidermolysis bullosa (Jonkman and Pasmooij, 2009), respectively. "
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    • "This transversion was assumed to be mediated by methylation. In addition, Wada et al. (2003, 2004) implied that DNA polymerase slippage was the cause of the compensatory deletions in two of their families . In one family, the function of the WAS protein was restored in two brothers by the same second-site mutation , a deletion of 19 bp that included the original 1305insC mutation in the WASP gene (Wada et al. Figure 6 Multiple reversion mutations in patients 1 and 2. In mitosis in a diploid cell after DNA replication and normal segregation, each daughter cell obtains one chromosome from the father and one from the mother. "
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    ABSTRACT: Revertant mosaicism by somatic reversion of inherited mutations has been described for a number of genetic diseases. Several mechanisms can underlie this reversion process, such as gene conversion, crossing-over, true back mutation, and second-site mutation. Here, we report the occurrence of multiple corrections in two unrelated probands with revertant mosaicism of non-Herlitz junctional epidermolysis bullosa, an autosomal recessive genodermatosis due to mutations in the COL17A1 gene. Immunofluorescence microscopy and laser dissection microscopy, followed by DNA and RNA analysis, were performed on skin biopsy specimens. In patient 1, a true back mutation, 3781T-->C, was identified in the specimen from the arm, and a second-site mutation, 4463-1G-->A, which compensated for the frameshift caused by the inherited 4424-5insC mutation, was identified in the 3' splice site of exon 55 in a specimen from the middle finger. Patient 2 showed--besides two distinct gene conversion events in specimens from the arm and hand sites, both of which corrected the 1706delA mutation--a second-site mutation (3782G-->C) in an ankle specimen, which prevented the premature ending of the protein by the 3781C-->T nonsense mutation (R1226X). Thus, both inherited mutations, paternal as well as maternal, reverted at least once by different reversion events in distinct cell clusters in the described patients. The occurrence of multiple correcting mutations within the same patient indicates that in vivo reversion is less unusual than was generally thought. Furthermore, in the male patient, mosaic patterns of type XVII collagen-positive keratinocytes were present in clinically unaffected and affected skin. This latter observation makes it likely that reversion may be overlooked and may happen more often than expected.
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