IMEF SDHC-loss ChIP-seq
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE104362 | GEO | 2018/02/15
REPOSITORIES: GEO
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Project description:We have developed a tet-inducible immortalized mouse embryonic fibroblast (iMEF) cell culture model of SDH-loss paraganglioma/pheochromocytoma in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). Using this model and an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt), we have characterized the time-course of Sdhc gene rearrangement, protein loss, and succinate accumulation following induction with doxycycline. Using RNA-seq, we have quantified changes in gene expression due to succinate accumulation using our Sdhc -/- cell line. Through a time-course experimental design, we have grown R26M2rtTA/+;TetOcre;Sdhcfl/fl (experimental) and R26M2rtTA/+;TetOcre;Sdhcfl/wt (control) iMEFs in standard DMEM media containing 10% FBS and quantified transcriptional changes in these cells as a function of time after triggering SDHC gene rearrangement via doxycycline exposure.
2017-11-21 | GSE103662 | GEO
Project description:We have developed a tet-inducible immortalized mouse embryonic fibroblast (iMEF) cell culture model of SDH-loss paraganglioma/pheochromocytoma in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). Using this model and an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt), we have characterized the time-course of Sdhc gene rearrangement, protein loss, and succinate accumulation following induction with doxycycline. Using reduced representation bisulfite sequencing (RRBS), we have quantified changes in the genome-wide distribution of cytosine/5’-methyl-cytosine due to succinate accumulation using our Sdhc -/- cell line. Through a time-course experimental design, we have grown R26M2rtTA/+;TetOcre;Sdhcfl/fl (experimental) and R26M2rtTA/+;TetOcre;Sdhcfl/wt (control) iMEFs in standard DMEM media containing 10% FBS and quantified DNA methylation changes in these cells as a function of time after triggering SDHC gene rearrangement via doxycycline exposure.
2017-11-21 | GSE103609 | GEO
Project description:We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure. Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, chromatin epigenomic marks including H3K4me3, H3K27me2, H3K27ac, CTCF, acetyllysine, propionyllysine, and butyryllysine were characterized by CUT&RUN.
2020-10-22 | GSE129954 | GEO
Project description:We have developed a tet-inducible SV-40 lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of SDH-loss paraganglioma/pheochromocytoma in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure. Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (experimental) and Sdhc +/- (control) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, transcriptomic profiling for each cell line was performed by RNA-seq.
2018-05-10 | GSE114244 | GEO
Project description:We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure. Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, long-range genomic contacts were profiled using eHi-C.
2020-10-22 | GSE129955 | GEO
Project description:We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure. Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, accessible chromatin regions in each cell line were interrogated by ATAC-seq.
2020-10-22 | GSE129953 | GEO
Project description:We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). Following doxycycline induction, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, cells were treated with histone acetyltransferase inhibitors C646 (10 µM) and MB-3 (100 µM) or vehicle for 3 days. Accessible chromatin regions were subsequently interrogated by ATAC-seq.
2020-10-22 | GSE145754 | GEO
Project description:iMEF SDHC-loss stable line CUT&RUN
| PRJNA533320 | ENA
Project description:iMEF SDHC-loss stable line ATAC-seq
| PRJNA533325 | ENA