title: [A study on alterations of gene expression in a flat revertant R1 from ras-oncogene transformed NIH/3T3 cells].
abstract: The flat revertant cell line R1, isolated from human activated Ha-ras oncogene transformed NIH/3T3 cells (EJ-NIH/3T3) by mutagen treatment, expresses a variant form of the actin-regulatory protein gelsolin, designated p92-5.7. To clone the gene encoding p92-5.7, gelsolin cDNAs were isolated from a cDNA library of R1 cells. In vitro transcription-translation and nucleotide sequence analyses of the cloned cDNAs identified a point mutation in codon 321 at the cause for the expression of p92-5.7. Considering gelsolin's function as an actin binding protein, the expression of alpha-actin, which is downregulated in many transformed fibroblasts, was analyzed. In EJ-NIH/3T3 cells no alpha-actin transcript was detected, whereas in R1 cells alpha-actin mRNA expression was restored to a level similar to NIH/3T3 cells. Immunofluorescence staining of the cells with an alpha-actin specific monoclonal antibody did not detect any alpha-actin containing microfilaments in EJ-NIH/3T3 cells, but revealed an ordered microfilament pattern in R1 and NIH/3T3 cells. In order to identify other genetic alterations that may also contribute to the revertant phenotype, genes with an elevated expression in R1 cells compared with the parental EJ-NIH/3T3 cells were isolated by using a differential hybridization approach. The identified sequences represented mitochondrial (cytochrome b, cytochrome c oxidase subunit II, NADH dehydrogenase subunits 1 and 4) and alpha 2 (type I) collagen genes. In summary, these results suggest that a complex alteration of the expression of cytoskeletal, mitochondrial and extracellular matrix components is closely associated with the flat reversion of R1 cells.
authors.affiliation: Laboratory of Molecular Genetics, Hokkaido University School of Medicine, Sapporo, Japan.
authors.forename: L
authors.initials: L
authors.lastname: Müllauer
mesh_terms: D016475:3T3 Cells / Q000502:physiology*; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis / Q000138:chemical synthesis; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis* / Q000235:genetics; D018260:Gelsolin / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology*; D011905:Genes, ras; D006801:Humans; D051379:Mice; D008840:Microfilament Proteins / Q000096:biosynthesis* / Q000235:genetics; D008856:Microscopy, Fluorescence; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000032:analysis
chemical_list: D018076:DNA, Complementary; D016326:Extracellular Matrix Proteins; D018260:Gelsolin; D008840:Microfilament Proteins; D012313:RNA