Mosaic and Intronic Mutations in TSC1/TSC2 Explain the Majority of TSC Patients with No Mutation Identified by Conventional Testing

Abstract

Tuberous sclerosis complex (TSC) is an autosomal dominant tumor suppressor gene syndrome due to germline mutations in either TSC1 or TSC2. 10–15% of TSC individuals have no mutation identified (NMI) after thorough conventional molecular diagnostic assessment. 53 TSC subjects who were NMI were studied using next generation sequencing to search for mutations in these genes. Blood/saliva DNA including parental samples were available from all subjects, and skin tumor biopsy DNA was available from six subjects. We identified mutations in 45 of 53 subjects (85%). Mosaicism was observed in the majority (26 of 45, 58%), and intronic mutations were also unusually common, seen in 18 of 45 subjects (40%). Seventeen (38%) mutations were seen at an allele frequency < 5%, five at an allele frequency < 1%, and two were identified in skin tumor biopsies only, and were not seen at appreciable frequency in blood or saliva DNA. These findings illuminate the extent of mosaicism in TSC, indicate the importance of full gene coverage and next generation sequencing for mutation detection, show that analysis of TSC-related tumors can increase the mutation detection rate, indicate that it is not likely that a third TSC gene exists, and enable provision of genetic counseling to the substantial population of TSC individuals who are currently NMI.

Author Summary

Tuberous sclerosis complex (TSC) is a human genetic disorder due to mutations in the TSC1 or TSC2 genes. A mystery for many years has been the fact that with standard genetic testing 10–15% of TSC patients have had no mutation identified (NMI) in either TSC1 or TSC2. We examined the genetic cause of TSC in patients who were ‘NMI’ after previous testing. We found a mutation in TSC1 or TSC2 in the vast majority of the subjects studied: 45 of 53 (85%). The majority of mutations identified were either in introns or mosaic or both. Usually we expect to find mutations causing human disease in exons, coding parts of genes. However, mutations can also be found in introns, the non-coding parts of genes, and we found intronic mutations in 18 of 45 subjects (40%). Mosaic mutations were seen in 26 of 45 subjects (58%). Mosaicism is the situation in which different cells in the body have a different genetic make-up, and in this case the mutations in TSC1/TSC2 were present in only a fraction of the cells from the patient. So these two types of hard-to-find mutations (in introns and/or mosaic) explain the majority of TSC patients who were NMI.