Linkage of the Wiskott-Aldrich syndrome with polymorphic DNA sequences from the human X chromosome.

Abstract

The The immune defects in WAS are poorly understood but include depressed serum IgM, elevated IgA and IgE, lack of response to polysaccharide antigens and thus absent isohaemagglutinins, reduced T-cell functions such as delayed hypersensitivity and graft rejection, and production of low numbers of abnormally small platelets that are rapidly eliminated, resulting in thrombocytopenia (5, 6). Despite extensive research efforts, the underlying biochemical defect and molecular basis of WAS remain unknown. Selective inactivation of the X chromosome carrying the WAS gene renders the heterozygous females serologically and clinically normal, thereby precluding detection of the carrier state (7). Thus, no direct test is available for prenatal diagnosis or carrier screening.In the absence of information on the primary product of a gene, one approach to locating and eventually cloning the gene is to use restriction fragment length polymorphic (RFLP) markers in conjunction with linkage analysis to study families in which the disease segregates.As a first step in providing diagnostic information for screening and prenatal diagnosis, and in identifying the primary molecular defect in WAS, we have used genetic linkage analysis with a series of X chromosome-specific cloned DNA DNA Extraction and Analysis. Total genomic DNA was extracted from peripheral blood or lymphoblastoid cell lines. Ten micrograms ofDNA was digested to completion with the appropriate restriction endonucleases, size-fractionated over 1% agarose gels, and transferred by the method of Southern to nylon membranes (8).Probes. Six X-chromosome-specific cloned DNA probes, corresponding to loci DXYS1, DXS1, DXS14, DXS7, DXS84, and OTC, were used in the linkage analysis (9)(10)(11)(12)(13)(14)(15)(16)(17). As shown in Table 1, each probe detects two allelic fragments, with the exception of probe pHO731, which defines a four-allele Msp I polymorphism at the ornithine transcarbamoylase locus (16). For hybridization, the probes were radiolabeled by random priming to a specific activity of 2 x 101 cpm/pgg (18

Identification of WASP mutations in 14 Spanish families with Wiskott-Aldrich syndrome

“…The gene for WAS was located to the short arm of the X chromosome at Xp11.22-p11.23 [Peacocke and Siminovitch, 1987;Kwan et al, 1991] and has been identi®ed by positional cloning [Derry et al, 1994]. This gene, designated WASP, is composed of 12 exons spanning 9 kb of genomic DNA; it encodes a protein of 502 amino acids.…”

Genetic study of a new X-linked recessive immunodeficiency syndrome.

“…XL lymphoproliferative syndrome is characterized by an abnormal susceptibility to Epstein-Barr virus (EBV) infection, the gene being localized in Xq25 (6). Finally, Wiskott-Aldrich syndrome (WAS) is associated with platelet anomalies and has been mapped around Xpl 1.2 (7,8).…”

A novel X-linked combined immunodeficiency disease.

“…and led to investigations of the role of the X chromosome in the genesis of that system. Genes for each of those X-linked immunologic deficiencies were mapped to specific regions of the X chromosome (19)(20)(21)(22)(23)(24)(25)(26)(27)(28). Furthermore, in X-linked CGD the resultant protein abnormality, a deficiency in the production of the 90-kD carrier protein for cytochrome b245, was elucidated (29, 30), and a sialic acid rich glycoprotein (GP-1 15) was found to be deficient in certain cases of WAS (31).…”