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Gene expression profiling on C3H/10T1/2 fibroblasts during their myogenic conversion in case of Six4 ablation

ABSTRACT: In this study, we used C3H/10T1/2 cells, a well known model of myogenic conversion, to study the effect of Six4 knockdown on the expression of genes during fibroblasts to myocytes conversion induced by ectopic expression of MyoD

ORGANISM(S): Mus musculus

PROVIDER: GSE66319 | GEO | 2015/12/31

SECONDARY ACCESSION(S): PRJNA276526

REPOSITORIES: GEO

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Project description:The two-stage cell transformation assay is an in vitro model cell culture system to identify the ability of chemicals to act as initiators or promoters of cell transforma- tion and also to study the cellular and molecular mechanisms of chemically induced morphological and neoplastic cell transformation. The global gene expression profiles of 3- methylcholanthrene (MCA)+12-O-tetradecanoylphorbol-13- acetate (TPA)-transformed C3H/10T1/2 cells are not known. Therefore, we have investigated the global transcriptional profile of MCA+TPA-transformed C3H10T1/2 cells using an 8M-CM-^W60 k probe microarray. The study revealed a differential regulation of pathways and gene expressions. Multifold dysregulation was seen in pathways of cancer, phagosomal activity, and tumor cell microenvironment information pro- cessing systems, notably the neuroactive ligandM-bM-^@M-^Sreceptor in- teraction, actin cytoskeleton regulation, tight junction, axon guidance, and cell adhesion molecules. The genes FGF1, EIF4E1B, MAGI1,and GRIA3 showed upregulation; these encoded the pluripotent fibroblast growth factor, the transla- tion initiation factor, the tight junction scaffolding protein, and the antiapoptotic as well as the enhancer of proliferation and migration, respectively. The genes CXCL7/CXCL5/CXCL12, H2DMB1,and HSPA1A showed downregulation; these encoded the chemotactic agent protein, the protein involved Total mRNA was isolated from the control as well as the MCA+TPA-transformed C3H/10T1/2 cells and complemen- tary RNA (cRNA) was prepared from mRNA (1 M-NM-<g). One- color microarray processing was performed. Acceptable qual- ity of the total RNA sample was ascertained by its electropho- resis trace and integrity assay using a Bioanalyzer which profiled RNA by RIN interpretation. The T7 promoter-based linear amplification was used to generate labeled complemen- tary RNA to amplify target material and incorporate cyanine 3-labeled CTP using AgilentM-bM-^@M-^Ys low-input RNA linear amplifi- cation kit one color (cat no. 5188-5339). The fluorescence- labeled cRNA samples were hybridized onto a Genotypic- designed Custom Whole Genome Mouse 8x60k slide (AMADID no. 26986) in duplicate using AgilentM-bM-^@M-^Ysinsitu hybridization kit (no. 5184-3568). Hybridization was carried out in AgilentM-bM-^@M-^Ys Surehyb Chambers at 65M-BM-0C for 16 h. The hybridized slides were washed using Agilent Gene Expression wash buffers (part no. 5188-5327). Fluorescence data were collected using an Agilent Microarray Scanner (G2567AA) and analyzed with program Gene Spring GX, version 11.5 (Agilent Technologies, Bangalore, India). Significantly dysregulated genes were identified. Statistical t test p value was calculated based on volcano plot using Gene spring GX Software.

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Gene expression profiling on C3H/10T1/2 fibroblasts during their myogenic conversion in case of Six4 ablation

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