Tuberous sclerosis complex (TSC) is a dominantly inherited disorder of hamartomas and hamartia with an estimated population frequency of 1 in 10,000. Both hamartomas and hamartia represent developmental anomalies characterized by abnormal cell or tissue arrangement and defective function. Hamartomas are distinguished by the additional feature of excessive cellular proliferation. The hamartomas do not usually progress to malignancy but cause morbidity and mortality due to size and location. Nearly every organ system has been reported to have the tumors in affected individuals; however, the most severe effects are in the CNS. Primary sources of morbidity are seizures (80-90% of affected individuals), mental retardation (40-50% of affected individuals) and behavioral abnormalities (most prominently autism; up to 50% of affected individuals). Efforts to clone the responsible genes have been extensive. Through genetic linkage studies, we know that there are two genes (on chromosomes 9q34 and 16p13) which probably account for all familial cases. The goal of this project is to clone the TSC gene on chromosome 9q34. Through genetic mapping studies, the critical TSC region on 9q34 has been narrowed to a genetic distance of two centimorgans and a physical region of less than two metabases. The project is moving from the genetic mapping phase into the physical mapping phase. Initially, I will continue genetic mapping studies by finding new families with multiple mapping phase. Initially, I will continue genetic mapping studies by finding new families with multiple affected individuals to identify additional key recombinants. Yeast artificial chromosome (YACs) which cover the critical region are being obtained from YAC libraries and by a new marker-specific method which is described. Candidate genes (by location) will be isolated from the region by Alu-PCR, zoo blotting, exon trapping and a novel in-vitro splicing technique. Candidate genes (by function) will be sought by screening the region for trinucleotide repeat sequences and screening the YACs at low stringency for functional motifs such as those know to serve in cell cycle regulation. Mutations to define the TSC gene will be searched for by: Southern blotting searching for large deletions, insertions and rearrangements; sequencing of candidates cDNAs to perform SSCP seeking point mutations and small deletions or insertions; and sequencing of DNAs from affected individuals in the event that the other two deletions or insertion; and sequencing of DNAs from affected individuals in the event that the other two strategies are unsuccessful. Once found, the TSC gene will be compared in normal and affected individuals to uncover more information about the disease and facilitate diagnosis and management.
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