Journal: Neoplasia (New York, N.Y.)

Article Title: Global Reduction of H3K4me3 Improves Chemotherapeutic Efficacy for Pediatric Ependymomas 1

doi: 10.1016/j.neo.2019.03.012

Figure Lengend Snippet: Candidate genes potentially associated with chemo- and radio-therapeutic resistance and histone H3K4me3 modification, identified by comparison of EPN samples with normal tissue from four-ventricular regions.

Article Snippet: Human His-tagged SETD1A expression plasmid, pET28-SETD1A-MHL, and its control pET28-MHL were gifts from Cheryl Arrowsmith (Addgene plasmid # 32868 and #26096, respectively).

Techniques: Modification, Binding Assay, Derivative Assay

Journal: Neoplasia (New York, N.Y.)

Article Title: Global Reduction of H3K4me3 Improves Chemotherapeutic Efficacy for Pediatric Ependymomas 1

doi: 10.1016/j.neo.2019.03.012

Figure Lengend Snippet: Reduction of H3K4me3 via down-regulation of hSETD1A decreases ERBB2 and CCND1 expression and increases chemosensitivity. A, Western blots showing hSETD1A, ERBB2, CCND1 and H3K4me3 in pediatric primary cultured cell lines. B and C, Real-time PCR (B) and western blots (C) show increased ERBB2 and CCND1 expression and H3K4me3 levels in pediatric primary cultured cells following treatment with pET28-SETD1A-MHL (pET-SETD1A), compared to pET28-MHL (pET28) control. D, Chromatin immunoprecipitation with H3K4me3 IgG antibodies coupled with real-time PCR (ChIP-PCR) using CCND1 and ERBB2 promoter primers in pediatric primary cultured EPN cells following treatment with pET-SETD1A, compared to pET28. E, MTS assays showing cell viability in response to CPL or VCR following treatment with pET-SETD1A or pET28 in pediatric primary EPN cells. F and G, Real-time PCR (F) and western blots (G) show decreased ERBB2 and CCND1 expression and H3K4me3 levels in pediatric primary cultured cells following treatment with siSETD1A (pET-SETD1A), compared to control siRNA (siCtrl). D, Chromatin immunoprecipitation with H3K4me3 IgG antibodies coupled with real-time PCR (ChIP-PCR) using CCND1 and ERBB2 promoter primers in pediatric primary cultured EPN cells following treatment with siSETD1A and siCtrl. I, MTS assays showing cell viability in response to CPL or VCR following treatment with siCtrl or siSETD1A in pediatric primary EPN cells. Error bars show the standard deviation of three independent experiments. * P < .05.

Article Snippet: Human His-tagged SETD1A expression plasmid, pET28-SETD1A-MHL, and its control pET28-MHL were gifts from Cheryl Arrowsmith (Addgene plasmid # 32868 and #26096, respectively).

Techniques: Expressing, Western Blot, Cell Culture, Real-time Polymerase Chain Reaction, Chromatin Immunoprecipitation, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: In Vivo and In Vitro Characterization of the RNA Binding Capacity of SETD1A (KMT2F)

doi: 10.3390/ijms242216032

Figure Lengend Snippet: Analysis of the mRNAs associated with SETD1A. ( A ). Number of mRNAs that showed higher FPKM values than the negative control in the three independent RIP experiments. ( B ). RT-qPCR results of the selected mRNAs. Fold enrichment of immunoprecipitated RNAs was calculated as 2 (−ΔΔCt [SETD1A RIP/NS Rb IgG RIP]) . Mean values and standard errors of three independent biological replicas are shown. Statistical analysis was performed using one-way ANOVA; *, **, and *** indicate significant differences from non-specific rabbit IgG control samples at p < 0.05, p < 0.005, and p < 0.001 values, respectively. ( C ). PANTHER protein classes that were significantly enriched in the mRNAs present in all three RIP-seq datasets (FDR adjusted p -value < 0.05). Functional classes related to RNA transcription regulation and RNA processing are labeled green and the DNA damage response class is labeled light blue. ( D ). IGV views of the mapped NGS reads for the selected genes: KDM4D (Lysine-specific demethylase 4D), SRPK2 (SRSF protein kinase 2), TET2 (Methylcytosine dioxygenase TET2), and SETD1B (Histone-lysine N-methyltransferase SETD1B). Reads in the NGS data are represented by gray columns, and intronic regions are highlighted by red bars. Exons are represented by blue boxes.

Article Snippet: DNA sequence coding for SETD1A_RRM (89-197, UniProt: O15047) domain in pET28-MHL vector was a gift from Cheryl Arrowsmith (Addgene plasmid #32868; http://n2t.net/addgene:32868 ; accessed on 12 April 2021; RRID: Addgene_32868).

Techniques: Negative Control, Quantitative RT-PCR, Immunoprecipitation, Control, Functional Assay, Labeling

Journal: International Journal of Molecular Sciences

Article Title: In Vivo and In Vitro Characterization of the RNA Binding Capacity of SETD1A (KMT2F)

doi: 10.3390/ijms242216032

Figure Lengend Snippet: Analysis of the lncRNAs associated with SETD1A. ( A ). Number of lncRNAs that showed higher FPKM values than the negative control in the three independent RIP experiments. ( B ). RT-qPCR results of the selected lncRNAs. Fold enrichment of immunoprecipitated RNAs was calculated as 2 (−ΔΔCt [SETD1A RIP/NS Rb IgG RIP]) . Mean values and standard errors of three independent biological replicas are shown. Statistical analysis was performed using one-way ANOVA; *, **, and **** indicate significant differences from non-specific rabbit IgG control samples at p < 0.05, p < 0.005, and p < 0.0001 values, respectively. ( C ). IGV views of the mapped NGS reads for the selected genes: LINC01521, SNHG8 (Small Nuclear Host Gene 8), TP53TG1 (TP53 Target 1), OLMALINC (Oligodendrocyte Maturation-associated Long Intergenic Non-coding RNA), HOTAIR (HOX Transcript Antisense RNA), and NORAD (Non-coding RNA Activated by DNA Damage). Reads in the NGS data are represented by gray columns, and intronic regions are highlighted by red bars. Exons are represented by blue boxes.

Article Snippet: DNA sequence coding for SETD1A_RRM (89-197, UniProt: O15047) domain in pET28-MHL vector was a gift from Cheryl Arrowsmith (Addgene plasmid #32868; http://n2t.net/addgene:32868 ; accessed on 12 April 2021; RRID: Addgene_32868).

Techniques: Negative Control, Quantitative RT-PCR, Immunoprecipitation, Control