title: Novel mutations of the APC gene and genetic consequences of splicing mutations in the Czech FAP families.
abstract: Familial adenomatous polyposis (FAP) is an autosomal dominant syndrome with almost 100 % risk of colorectal cancer. The typical FAP is characterized by hundreds to thousands of colorectal adenomatous polyps and by extracolonic manifestations, later onset and lower number of polyps in colon is characteristic of an attenuated form (AFAP). We analyzed the APC gene for germline mutations in 90 FAP/AFAP patients. Mutation screening was performed using Denaturing Gradient Gel Electrophoresis. DNA fragments showing an aberrant electrophoretic banding pattern were sequenced. APC-mutation-negative probands were screened for large deletions of the APC gene using multiplex ligation dependent probe amplification. Analysis of mRNA variants followed in probands with possible splicing mutation by PCR amplification of target site flanking exons and sequencing the normal and aberrant products. We identified 30 germline variants among 36 unrelated probands including large deletions. Eleven APC variants detected last two years have not been reported yet. At all, fifteen of them are expected to cause errors in mRNA splicing. Analysis of mRNA in ten of these patients revealed exon skipping in seven cases, exonisation of intron in one of these as well, change of the amount of alternatively spliced product in one case, and no effect was found in three cases. In two of the patients, the biopsy of colon mucosa and polyp enabled us to examine the effect of the mutation on splicing pattern in colon cells directly. The comparison of alternative and standard transcript amount showed similar transcription pattern of exon 14 in control colon mucosa tissue (9 samples) as in 51 blood control samples.
authors.affiliation: Institute of Biology and Medical Genetics of the 1st Faculty of Medicine and General Teaching Hospital, Charles University in Prague, Albertov 4, Prague 2, 128 00, Czech Republic. lucie.schwarzova@yahoo.com
authors.forename: Lucie
authors.initials: L
authors.lastname: Schwarzová
authors.forename: Jitka
authors.initials: J
authors.lastname: Štekrová
authors.forename: Martina
authors.initials: M
authors.lastname: Florianová
authors.forename: Aleš
authors.initials: A
authors.lastname: Novotný
authors.forename: Michaela
authors.initials: M
authors.lastname: Schneiderová
authors.forename: Petr
authors.initials: P
authors.lastname: Lněnička
authors.forename: Věra
authors.initials: V
authors.lastname: Kebrdlová
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Kotlas
authors.forename: Kamila
authors.initials: K
authors.lastname: Veselá
authors.forename: Milada
authors.initials: M
authors.lastname: Kohoutová
mesh_terms: D011125:Adenomatous Polyposis Coli / Q000235:genetics*; D017398:Alternative Splicing; D015179:Colorectal Neoplasms / Q000235:genetics*; D018153:Czech Republic; D005260:Female; D017491:Genes, APC; D020022:Genetic Predisposition to Disease; D006801:Humans; D008297:Male; D009154:Mutation