title: X-linked Alport syndrome caused by splicing mutations in COL4A5.
abstract: OBJECTIVE
  X-linked Alport syndrome is caused by mutations in the COL4A5 gene. Although many COL4A5 mutations have been detected, the mutation detection rate has been unsatisfactory. Some men with X-linked Alport syndrome show a relatively mild phenotype, but molecular basis investigations have rarely been conducted to clarify the underlying mechanism.
  
  
  METHODS
  In total, 152 patients with X-linked Alport syndrome who were suspected of having Alport syndrome through clinical and pathologic investigations and referred to the hospital for mutational analysis between January of 2006 and January of 2013 were genetically diagnosed. Among those patients, 22 patients had suspected splice site mutations. Transcripts are routinely examined when suspected splice site mutations for abnormal transcripts are detected; 11 of them showed expected exon skipping, but others showed aberrant splicing patterns. The mutation detection strategy had two steps: (1) genomic DNA analysis using PCR and direct sequencing and (2) mRNA analysis using RT-PCR to detect RNA processing abnormalities.
  
  
  RESULTS
  Six splicing consensus site mutations resulting in aberrant splicing patterns, one exonic mutation leading to exon skipping, and four deep intronic mutations producing cryptic splice site activation were identified. Interestingly, one case produced a cryptic splice site with a single nucleotide substitution in the deep intron that led to intronic exonization containing a stop codon; however, the patient showed a clearly milder phenotype for X-linked Alport syndrome in men with a truncating mutation. mRNA extracted from the kidney showed both normal and abnormal transcripts, with the normal transcript resulting in the milder phenotype. This novel mechanism leads to mild clinical characteristics.
  
  
  CONCLUSIONS
  This report highlights the importance of analyzing transcripts to enhance the mutation detection rate and provides insight into genotype-phenotype correlations. This approach can clarify the cause of atypically mild phenotypes in X-linked Alport syndrome.
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan; nozu@med.kobe-u.ac.jp.
authors.forename: Kandai
authors.initials: K
authors.lastname: Nozu
authors.affiliation: Faculty of Medicine, University of Southampton, Southampton, United Kingdom;
authors.forename: Igor
authors.initials: I
authors.lastname: Vorechovsky
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan;
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kaito
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan;
authors.forename: Xue Jun
authors.initials: XJ
authors.lastname: Fu
authors.affiliation: Department of Pediatrics, Wakayama Medical University, Wakayama, Japan;
authors.forename: Koichi
authors.initials: K
authors.lastname: Nakanishi
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan;
authors.forename: Yuya
authors.initials: Y
authors.lastname: Hashimura
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan;
authors.forename: Fusako
authors.initials: F
authors.lastname: Hashimoto
authors.affiliation: Division of Nephrology and Rheumatology, National Center for Child Health and Development, Tokyo, Japan;
authors.forename: Koichi
authors.initials: K
authors.lastname: Kamei
authors.affiliation: Division of Nephrology and Rheumatology, National Center for Child Health and Development, Tokyo, Japan;
authors.forename: Shuichi
authors.initials: S
authors.lastname: Ito
authors.affiliation: Department of Nephrology, Fukuoka Children's Hospital, Fukuoka, Japan;
authors.forename: Yoshitsugu
authors.initials: Y
authors.lastname: Kaku
authors.affiliation: Kidney Center, National Hospital Organization Chiba-East Hospital, Chiba, Japan;
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Imasawa
authors.affiliation: Department of Pediatrics, Yokkaichi Municipal Hospital, Mie, Japan;
authors.forename: Katsumi
authors.initials: K
authors.lastname: Ushijima
authors.affiliation: Department of Pediatrics, National Hospital Organization Okayama Medical Center, Okayama, Japan;
authors.forename: Junya
authors.initials: J
authors.lastname: Shimizu
authors.affiliation: Education Center, Asahikawa Medical University, Hokkaido, Japan; and.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Makita
authors.affiliation: Department of Pediatrics, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Konomoto
authors.affiliation: Department of Pediatrics, Wakayama Medical University, Wakayama, Japan;
authors.forename: Norishige
authors.initials: N
authors.lastname: Yoshikawa
authors.affiliation: Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan;
authors.forename: Kazumoto
authors.initials: K
authors.lastname: Iijima
mesh_terms: D000293:Adolescent; D000328:Adult; D002648:Child; D002675:Child, Preschool; D024141:Collagen Type IV / Q000235:genetics*; D004252:DNA Mutational Analysis; D005091:Exons; D005260:Female; D005838:Genotype; D006801:Humans; D007438:Introns; D008297:Male; D008875:Middle Aged; D009154:Mutation; D009394:Nephritis, Hereditary / Q000235:genetics*; D010641:Phenotype; D022821:RNA Splice Sites; D012333:RNA, Messenger / Q000032:analysis*; D055815:Young Adult
chemical_list: C478159:COL4A5 protein, human; D024141:Collagen Type IV; D022821:RNA Splice Sites; D012333:RNA, Messenger
keywords: Alport syndrome; genetic renal disease; glomerular disease