title: The RET p.G533C mutation confers predisposition to multiple endocrine neoplasia type 2A in a Brazilian kindred and is able to induce a malignant phenotype in vitro and in vivo.
abstract: BACKGROUND
  We have previously described a p.G533C substitution in the rearranged during transfection (RET) oncogene in a large family with medullary thyroid carcinoma. Here, we explore the functional transforming potential of RET p.G533C mutation.
  
  
  METHODS
  Plasmids expressing RET mutants (p.G533C and p.C634Y) and RET wild type were stable transfected into a rat thyroid cell line (PCCL3). Biological and biochemical effects of RET p.G533C were investigated both in vitro and in vivo. Moreover, we report the first case of pheochromocytoma among the RET p.G533C-carriers in this Brazilian family and explore the RET mutational status in DNA isolated from pheochromocytoma.
  
  
  RESULTS
  Ectopic expression of RET p.G533C and p.C634Y activates RET/MAPK/ERK pathway at similar levels and significantly increased cell proliferation, compared with RET wild type. We additionally show that p.G533C increased cell viability, anchorage-independent growth, and micronuclei formation while reducing apoptosis, hallmarks of the malignant phenotype. RET p.G533C down-regulates the expression of thyroid specific genes in PCCL3. Moreover, RET p.G533C-expressing cells were able to induce liver metastasis in nude mice. Finally, we described two novel RET variants (G548V and S556T) in the DNA isolated from pheochromocytoma while they were absent in the DNA isolated from blood.
  
  
  CONCLUSIONS
  Our in vitro and in vivo analysis indicates that this mutation confers a malignant phenotype to PCCL3 cells. These findings, in association with the report of first case of pheochromocytoma in the Brazilian kindred, suggest that this noncysteine mutation may be more aggressive than was initially considered.
authors.affiliation: Division of Genetics, Genetic Bases of Thyroid Tumors Laboratory, Federal University of São Paulo, São Paulo, Brazil.
authors.forename: Mariana N L
authors.initials: MN
authors.lastname: Oliveira
authors.forename: Jefferson P
authors.initials: JP
authors.lastname: Hemerly
authors.forename: André U
authors.initials: AU
authors.lastname: Bastos
authors.forename: Rosana
authors.initials: R
authors.lastname: Tamanaha
authors.forename: Flavia R M
authors.initials: FR
authors.lastname: Latini
authors.forename: Cléber P
authors.initials: CP
authors.lastname: Camacho
authors.forename: Anelise
authors.initials: A
authors.lastname: Impellizzeri
authors.forename: Rui M B
authors.initials: RM
authors.lastname: Maciel
authors.forename: Janete M
authors.initials: JM
authors.lastname: Cerutti
mesh_terms: D000293:Adolescent; D000310:Adrenal Gland Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D000704:Analysis of Variance; D000818:Animals; D017209:Apoptosis; D001938:Brazil; D018276:Carcinoma, Medullary / Q000151:congenital; D002448:Cell Adhesion; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology / Q000235:genetics / Q000556:secondary; D008297:Male; D051379:Mice; D008819:Mice, Nude; D048629:Micronuclei, Chromosome-Defective; D008875:Middle Aged; D018813:Multiple Endocrine Neoplasia Type 2a / Q000201:enzymology / Q000235:genetics* / Q000556:secondary; D009154:Mutation; D009386:Neoplastic Syndromes, Hereditary / Q000201:enzymology / Q000235:genetics*; D010375:Pedigree; D010641:Phenotype; D010673:Pheochromocytoma / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D010766:Phosphorylation; D051096:Proto-Oncogene Proteins c-ret / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D013961:Thyroid Gland / Q000201:enzymology* / Q000473:pathology; D013964:Thyroid Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000556:secondary; D014162:Transfection
chemical_list: D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human; D048049:Extracellular Signal-Regulated MAP Kinases