Wiskott-Aldrich syndrome/X-linked thrombocytopenia : WASP gene mutations, protein expression, and phenotype

• Published in: Blood 1997-10, Vol.90 (7), p.2680-2689
• Main Authors: ZHU, Q, WATANABE, C, LIU, T, HOLLENBAUGH, D, BLAESE, R. M, KANNER, S. B, ARUFFO, A, OCHS, H. D
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Biological and medical sciences; DNA Mutational Analysis; Exons - genetics; Genotype; Hematologic and hematopoietic diseases; Hematopoietic Stem Cells - metabolism; Humans; Medical sciences; Molecular Sequence Data; Phenotype; Platelet diseases and coagulopathies; Point Mutation; Protein Biosynthesis; Proteins - genetics; Rabbits; Sequence Deletion; Severity of Illness Index; Thrombocytopenia - genetics; Wiskott-Aldrich Syndrome - genetics; Wiskott-Aldrich Syndrome Protein; X Chromosome - genetics
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.v90.7.2680

Wiskott-Aldrich syndrome (WAS) and X-linked thrombocytopenia (XLT), caused by mutations of the WAS protein (WASP) gene, represent different phenotypes of the same disease. To demonstrate a phenotype/genotype correlation, we determined WASP gene mutations in 48 unrelated WAS families. Mutations included missense (20 families) and nonsense (eight) mutations located mostly in exons 1 to 4, and splice-site mutations (seven) and deletions and insertions (13) located preferentially in exons 7 to 11. Both genomic DNA and cDNA were sequenced and WASP expression was measured in cell lysates using peptide-specific rabbit anti-WASP antibodies. WASP was expressed in hematopoietic cell lines including bone marrow-derived CD34+ cells. Missense mutations located in exons 1 to 3 caused mild disease in all but one family and permitted WASP expression, although frequently at decreased concentration. Missense mutations affecting exon 4 were associated with classic WAS and, with one exception, barely detectable WASP. Nonsense mutations caused classic WAS and lack of protein. Insertions, deletions, and splice-site mutations resulted in classic WAS and absent, unstable, truncated, or multiply spliced protein. Using affinity precipitation, WASP was found to bind to Src SH3-containing proteins Fyn, Lck, PLC-gamma, and Grb2, and mutated WASP, if expressed, was able to bind to Fyn-glutathione S-transferase (GST) fusion protein. We conclude that missense mutations affecting the PH domain (exons 1 to 3) of WASP inhibit less important functions of the protein and result in a mild phenotype, and that missense mutations affecting exon 4 and complex mutations affecting the 3' portion of WASP interfere with crucial functions of the protein and cause classic WAS.