Mutation screening of the entire coding regions of the TSC1 and the TSC2 gene with the protein truncation test (PTT) identifies frequent splicing defects.

Mutation analyses in tuberous sclerosis (TSC) have reported a wide variety of disease-causing aberrations in the two known predisposing genes, TSC1 and TSC2 on chromosomes 9q34 and 16p13, comprising mainly small mutations distributed over the entire genes. So far, all known TSC1 mutations as well as the majority of TSC2 mutations truncate the proteins hamartin and tuberin, respectively. We describe for the first time an RNA-based screening of the entire coding regions of both TSC genes for truncating mutations applying the protein truncation test (PTT). Simultaneous investigation of both TSC genes in a group of 48 unassigned TSC patients, which were previously tested to exclude large intragenic TSC2 rearrangements, revealed aberrant migrating polypeptides resulting from truncating mutations in nine TSC1 cases and in 16 TSC2 cases while three TSC2 cases showed enlarged proteins. TSC1 mutations include two nonsense mutations, four insertions, and three splice mutations. Nineteen mutations identified in TSC2 were composed of four different nonsense mutations in five patients, one deletion, one insertion, and seven different splicing aberrations due to at least eight different mutations found in 12 patients. Additional predicted truncating mutations according to PTT without possible identification of the causative alteration allowed assignment to TSC1 in one and TSC2 in seven cases. Twelve patients without abnormalities in the PTT are assumed to harbor missense mutations, probably in TSC2. The high proportion of TSC2 splicing aberrations strengthens the importance of intronic disease-causing mutations and the application of RNA-based screening methods to confirm their consequences.