title: The methylation of the TSC2 promoter underlies the abnormal growth of TSC2 angiomyolipoma-derived smooth muscle cells.
abstract: Tuberous sclerosis complex (TSC) is an autosomal-dominant disease that is caused by mutations in either the TSC1 or TSC2 gene. Smooth muscle-like cells (ASMs) were isolated from an angiomyolipoma of a patient with TSC. These cells lacked tuberin, were labeled by both HMB45 and CD44v6 antibodies, and had constitutive S6 phosphorylation. The cells bear a germline TSC2 intron 8-exon 9 junction mutation, but DNA analysis and polymerase chain reaction amplification failed to demonstrate loss of heterozygosity. Testing for an epigenetic alteration, we detected methylation of the TSC2 promoter. Its biological relevance was confirmed by tuberin expression and a reduction in HMB45 labeling and S6 constitutive phosphorylation after exposure to the chromatin-remodeling agents, trichostatin A and 5-azacytidine. These cells were named TSC2(-/meth) ASMs. Their proliferation required epidermal growth factor in the medium as previously described for TSC2(-/-) ASMs. Blockade of epidermal growth factor with monoclonal antibodies caused the death of TSC2(-/meth) ASMs. In addition, rapamycin effectively blocked the proliferation of these cells. Our data show for the first time that methylation of the TSC2 promoter might cause a complete loss of tuberin in TSC2 cells, and that the pathogenesis of angiomyolipomas might also originate from epigenetic defects in smooth muscle cells. Additionally, the effect of chromatin-remodeling agents in these cells suggests a further avenue for the treatment of TSC as well as lymphangioleiomyomatosis.
authors.affiliation: Laboratory of Pharmacology, Dept. of Medicine, Surgery, and Dentistry, Via A. di Rudinì 8, 20142 Milano, Italy. elena.lesma@unimi.it
authors.forename: Elena
authors.initials: E
authors.lastname: Lesma
authors.forename: Silvia Maria
authors.initials: SM
authors.lastname: Sirchia
authors.forename: Silvia
authors.initials: S
authors.lastname: Ancona
authors.forename: Stephana
authors.initials: S
authors.lastname: Carelli
authors.forename: Silvano
authors.initials: S
authors.lastname: Bosari
authors.forename: Filippo
authors.initials: F
authors.lastname: Ghelma
authors.forename: Emanuele
authors.initials: E
authors.lastname: Montanari
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Di Giulio
authors.forename: Alfredo
authors.initials: A
authors.lastname: Gorio
mesh_terms: D000328:Adult; D018207:Angiomyolipoma / Q000209:etiology / Q000235:genetics* / Q000473:pathology*; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000951:Antigens, Neoplasm; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D019175:DNA Methylation / Q000235:genetics*; D004252:DNA Mutational Analysis; D018095:Germ-Line Mutation; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D007680:Kidney Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008297:Male; D058950:Melanoma-Specific Antigens; D008856:Microscopy, Fluorescence; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000473:pathology*; D009363:Neoplasm Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics; D020123:Sirolimus / Q000494:pharmacology; D014402:Tuberous Sclerosis / Q000150:complications / Q000235:genetics / Q000473:pathology; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: D000903:Antibiotics, Antineoplastic; D000951:Antigens, Neoplasm; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins; C000624653:TSC2 protein, human; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins; D020123:Sirolimus