title: Two-exon skipping due to a point mutation in p67-phox--deficient chronic granulomatous disease.
abstract: The cytosolic 67-kD protein in phagocytes (p67-phox) and B lymphocytes is one of essential components of the superoxide-generating system in these cells, and its defect causes an autosomal recessive type of chronic granulomatous disease (CGD). We performed mutation analysis of p67-phox mRNA from a CGD patient who lacks the protein and found an in-frame deletion from nucleotide 694 to 879, which corresponds to the entire sequence of exons 8 and 9. This sequence encodes one of two Src homology 3 domains and a part of proline-rich domain in p67-phox and lack of these domains seem to have influenced stability of this protein. To know causative reason for the deletion, we analyzed genomic DNA for p67-phox using two sets of primers that covered exons 8 and 9 with adjacent introns. The DNA fragments from the patient were shown to be same in length as those from control. However, the single-strand conformation-polymorphism analysis of the fragments showed that a patient's specimen that included the splice junction of exon 9 exhibited different mobility from the control. By sequencing of the fragment, a homozygous G to A replacement at position +1 of intron 9 was found to be a sole mutation, which reduced the matching score of the splicing sequence to the consensus calculated according to the formula proposed by Shapiro and Senapathy (Nucleic Acids Res 15:7155, 1987). The reduced matching score at the splice doner site (5' splice site) of intron 9 and the original low matching score at the acceptor site (3' splice site) of intron 7 may explain the skipping of exon 8 and 9, and another predicted mechanism is discussed on the basis of Shapiro and Senapathy's hypothesis.
authors.affiliation: Institute of Medical Science, University of Tokyo, Mitsubishi Yuka Bio-Clinical Laboratories Inc., Japan.
authors.forename: M
authors.initials: M
authors.lastname: Aoshima
authors.forename: H
authors.initials: H
authors.lastname: Nunoi
authors.forename: M
authors.initials: M
authors.lastname: Shimazu
authors.forename: S
authors.initials: S
authors.lastname: Shimizu
authors.forename: O
authors.initials: O
authors.lastname: Tatsuzawa
authors.forename: R T
authors.initials: RT
authors.lastname: Kenney
authors.forename: S
authors.initials: S
authors.lastname: Kanegasaki
mesh_terms: D000328:Adult; D001483:Base Sequence; D004252:DNA Mutational Analysis; D005091:Exons / Q000235:genetics*; D006105:Granulomatous Disease, Chronic / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000201:enzymology; D010750:Phosphoproteins / Q000172:deficiency / Q000235:genetics*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D012326:RNA Splicing / Q000235:genetics*; D018909:src Homology Domains
chemical_list: D010750:Phosphoproteins; C079789:neutrophil cytosol factor 67K