Inactivating mutations in an SH2 domain-encoding gene in X-linked lymphoproliferative syndrome

Abstract

X-linked lymphoproliferative syndrome (XLP) is an inherited immunodeficiency characterized by increased susceptibility to Epstein-Barr virus (EBV). In affected males, primary EBV infection leads to the uncontrolled proliferation of virus-containing B cells and reactive cytotoxic T cells, often culminating in the development of high-grade lymphoma. The XLP gene has been mapped to chromosome band Xq25 through linkage analysis and the discovery of patients harboring large constitutional genomic deletions. We describe here the presence of small deletions and intragenic mutations that specifically disrupt a gene named DSHP in 6 of 10 unrelated patients with XLP. This gene encodes a predicted protein of 128 amino acids composing a single SH2 domain with extensive homology to the SH2 domain of SHIP, an inositol polyphosphate 5-phosphatase that functions as a negative regulator of lymphocyte activation. DSHP is expressed in transformed T cell lines and is induced following in vitro activation of peripheral blood T lymphocytes. Expression of DSHP is restricted in vivo to lymphoid tissues, and RNA in situ hybridization demonstrates DSHP expression in activated T and B cell regions of reactive lymph nodes and in both T and B cell neoplasms. These observations confirm the identity of DSHP as the gene responsible for XLP, and suggest a role in the regulation of lymphocyte activation and proliferation. Induction of DSHP may sustain the immune response by interfering with SHIP-mediated inhibition of lymphocyte activation, while its inactivation in XLP patients results in a selective immunodeficiency to EBV.

Figure 1

Physical map of the XLP locus and identification of new genomic deletions disrupting DSHP. (A) Schematic representation of the XLP locus. The locus has been defined by a 3-Mb genomic deletion in patient A. Two new patients, R and AD, harbor deletions of 200 kb that span the DSHP gene, and are approximately 1 Mb centromeric to the previously reported 150 kb deletion in patient D. (B) Delineation of the borders of genomic deletions in patients R and AD, using PCR markers for centromeric and telomeric borders, and Southern blot analysis using probes from the 5′ and 3′ regions of DSHP. DSHP is entirely deleted in patient AD, whereas the centromeric breakpoint in patient R is between exons 1 and 2. (C) Comparison of the amino acid sequence of DSHP with the SH2 domain of SHIP. Identical amino acids are in black boxes, and conserved residues are in gray boxes. DSHP exon boundaries are indicated by arrowheads.

Figure 2

Northern blot analysis of DSHP expression. (A) DSHP expression in normal human tissues and in B, T, and myeloid cell lines, using a probe generated from the 3′ untranslated region. (B) Induction of DSHP expression following activation of PBLs using Con A or antibody to CD3.

Figure 3

DSHP expression in normal reactive lymph node and malignant lymphoma. (a) Strong expression of DSHP in some germinal center (g) and paracortical (p) cells as demonstated by RNA in situ hybridization using antisense probe. Cells of mantle zone (m) do not express DSHP. (Inset) High magnification of strongly expressing cells with activated morphologic features of vesicular nucleus and nucleolus. Hybridization using sense probe was negative. Immunohistochemical analysis of this section demonstrates (b) the distribution of B cells using antibody to the marker CD30 and (c) the distribution of T cells using antibody to CD3. (d) Strong expression of DSHP in atypical cells of diffuse large B cell lymphoma.