IDENTIFICATION OF NOVEL BRUTON-TYROSINE-KINASE MUTATIONS IN 10 UNRELATED SUBJECTS WITH X-LINKED AGAMMAGLOBULINEMIA

Mutations of the Bruton's tyrosine kinase (Btk) gene cause X linked ag ammaglobulinaemia (XLA). This inherited immunodeficiency disease cause s an arrest in B cell differentiation of pre-B cells to mature B cells . In this study we report the characterisation of mutations in the Btk gene in 10 unrelated XLA families. The screening approach we used was based on reverse transcriptase PCR and direct cycle sequencing of the amplified products followed by analysis of the observed mutations at the level of genomic DNA. The single strand confirmation polymorphism (SSCP) technique was used for assessment of the carriers in some of th ese families. Various mutations throughout the coding gene were observ ed, including missense and nonsense mutations, a deletion, and several splicing defects. None of the mutations except one has been previousl y described. There were three point mutations resulting in a single am ino acid substitution. One ofthese missense mutations was observed in a conserved region of the PH domain, the other two were found in the s rc homology domain 2 that is involved in phosphotyrosyl peptide bindin g. Two mutations were single base pair substitutions resulting in prem ature stop codons. In four patients abnormal Btk transcripts were foun d that were the result of aberrant splicing. One small deletion was ob served causing a frameshift and a secondary premature termination sign al. Characterisation of the mutations responsible for XLA allowed us t o diagnose the disease conclusively and identify the phenotypically no rmal female carriers.