Girls homozygous for an IL-2–inducible T cell kinase mutation that leads to protein deficiency develop fatal EBV-associated lymphoproliferation
Kirsten Huck, Oliver Feyen, Tim Niehues, Franz Rüschendorf, Norbert Hübner, Hans-Jürgen Laws, Tanja Telieps, Stefan Knapp, Hans-Heinrich Wacker, Alfons Meindl, Hassan Jumaa, Arndt Borkhardt
Kirsten Huck, Oliver Feyen, Tim Niehues, Franz Rüschendorf, Norbert Hübner, Hans-Jürgen Laws, Tanja Telieps, Stefan Knapp, Hans-Heinrich Wacker, Alfons Meindl, Hassan Jumaa, Arndt Borkhardt
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Research Article Immunology

Girls homozygous for an IL-2–inducible T cell kinase mutation that leads to protein deficiency develop fatal EBV-associated lymphoproliferation

Abstract

The fatal immune dysregulation that sometimes follows EBV infection in boys has been linked to mutations in two X chromosome–encoded genes, SLAM-associated protein (SAP) and X-linked inhibitor of apoptosis (XIAP). In this study we describe 2 girls from a consanguineous Turkish family who died after developing severe immune dysregulation and therapy-resistant EBV-positive B cell proliferation following EBV infection. SNP array–based genome-wide linkage analysis revealed IL-2–inducible T cell kinase (ITK) as a candidate gene for this immunodeficiency syndrome. Both girls harbored a homozygous missense mutation that led to substitution of a highly conserved residue (R335W) in the SH2 domain of ITK. Characteristics of ITK deficiency in mouse models, such as absence of NKT cells and high levels of eomesodermin in CD8+ cells, were seen in either one or both of the girls. Two lines of evidence suggested that R335W caused instability of the ITK protein. First, in silico modeling of the mutant protein predicted destabilization of the SH2 domain. Additionally, Western blot analysis revealed that, unlike wild-type ITK, the R335W mutant was nearly undetectable when expressed in 293 T cells. Our results suggest that ITK deficiency causes what we believe to be a novel immunodeficiency syndrome that leads to a fatal inadequate immune response to EBV. Because ITK deficiency resembles EBV-associated lymphoproliferative disorders in boys, we suggest that this molecular cause should be considered during diagnosis and treatment.

Authors

Kirsten Huck, Oliver Feyen, Tim Niehues, Franz Rüschendorf, Norbert Hübner, Hans-Jürgen Laws, Tanja Telieps, Stefan Knapp, Hans-Heinrich Wacker, Alfons Meindl, Hassan Jumaa, Arndt Borkhardt

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Figure 3

Evidence of severe destabilization of the mutant ITK protein.

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Evidence of severe destabilization of the mutant ITK protein. (A) Wester...
(A) Western blot with transiently transfected 293 T cells using a monoclonal ITK antibody. Cyclophilin A was used as negative control (lane 1). No expression was detected for the R335W mutant (lane 5) and an ITK variant with complete deletion of the SH2 domain (lane 3), whereas wild-type ITK (lane 4) and the kinase-inactivating mutant (F511Y) (lane 2) showed strong ITK expression. (B) Results of quantitative real-time PCR for EOMES mRNA in CD8+ single-positive T cells on a logarithmic scale. We found 1,000- to 6,000-fold higher mRNA levels of EOMES in both affected children compared with their heterozygous parents or healthy controls. Error bars denote the standard deviation. P1, patient 1; P2, patient 2; F, father; M, mother; C1, control 1; C2, control 2. (C) Domain architecture of ITK and the location of the mutated residue (R335) (highlighted by a red label) in the SH2 domain. Shown is the NMR minimized average structure (protein data bank code 2etz). Protein instability probably results from a compromised salt bridge network (dotted lines) as well as surface exposure of the introduced tryptophan side chain. A phosphotyrosine substrate peptide and the corresponding binding pocket are located on the opposite side of the molecule. The main structural elements are labeled. (D) Site of the BG loop mutations in ITK (R335W, red), BTK (K374N, blue), and SAP (Q99P, yellow).