title: Increased genomic instability and altered chromosomal protein phosphorylation timing in HRAS-transformed mouse fibroblasts.
abstract: The RAS-mitogen-activated protein kinase signaling pathway is often deregulated in cancer cells. In metastatic HRAS-transformed mouse fibroblasts (Ciras-3), the RAS-MAPK pathway is constitutively activated. We show here that Ciras-3 cells exhibit a higher incidence of chromosomal instability than 10T1/2 cells, including higher levels of clonal and nonclonal chromosomal aberrations. Stimulation of serum starved 10T1/2 and Ciras-3 cells with phorbol esters (TPA) results in the phosphorylation of histone H3 at serine 10 and serine 28. Regardless of the increased genomic instability in Ciras-3 cells, TPA-induced H3 phosphorylated at serine 10 and H3 phosphorylated at serine 28 partitioned into distinct nuclear subdomains as they did in the parental cells. However, the timing of the response of the H3 phosphorylation event to TPA induction was delayed in Ciras-3 cells. Further Ciras-3 cells, which have a more open chromatin structure, had increased steady state levels of phosphorylated H3 and HMGN1 relative to parental 10T1/2 cells. TPA-induced H3 phosphorylated at serine 10 and 28 were colocalized with the transcriptionally initiated form of RNA polymerase II in 10T1/2 and Ciras-3 cells. Chromatin immunoprecipitation assays demonstrated that TPA-induced H3 phosphorylation at serine 28 was associated with the immediate early JUN promoter, providing direct evidence that this histone post-translational modification is associated with transcriptionally active genes. Together our results demonstrate the increased genomic instability and alterations in the epigenetic program in HRAS-transformed cells.
authors.affiliation: Manitoba Institute of Cell Biology, University of Manitoba, Winnipeg, Manitoba, Canada R3E 0V9.
authors.forename: Katherine L
authors.initials: KL
authors.lastname: Dunn
authors.forename: Shihua
authors.initials: S
authors.lastname: He
authors.forename: Landon
authors.initials: L
authors.lastname: Wark
authors.forename: Geneviève P
authors.initials: GP
authors.lastname: Delcuve
authors.forename: Jian-Min
authors.initials: JM
authors.lastname: Sun
authors.forename: Hou
authors.initials: H
authors.lastname: Yu Chen
authors.forename: Sabine
authors.initials: S
authors.lastname: Mai
authors.forename: James R
authors.initials: JR
authors.lastname: Davie
mesh_terms: D000818:Animals; D015197:Carcinogenicity Tests; D002461:Cell Line, Transformed; D047369:Chromatin Immunoprecipitation; D002869:Chromosome Aberrations; D003627:Data Interpretation, Statistical; D005347:Fibroblasts; D015870:Gene Expression / Q000187:drug effects; D042822:Genomic Instability; D024241:HMGN1 Protein / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation; D015398:Signal Transduction; D032681:Spectral Karyotyping; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D024241:HMGN1 Protein; D006657:Histones; D010703:Phorbol Esters; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins