6a2be6a3c97a7ac3a0d47ceba4151763 270598.pdf ae869320b670692a25787929185563e2b870c43c 270598.pdf ff15401bc1137ee3f0231d0a573597354011932f461ddb2df19a16f898a65e8e 270598.pdf Title: The Histone Variant MacroH2A1 Impacts Circadian Gene Expression and Cell Phenotype in an In Vitro Model of Hepatocellular Carcinoma Subject: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. A foremost risk factor for HCC is obesity/metabolic syndrome-related non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH), which is prompted by remarkable changes in transcription patterns of genes enriching metabolic, immune/inflammatory, and circadian pathways. Epigenetic mechanisms play a role in NAFLD-associated HCC, and macroH2A1, a variant of histone H2A, is involved in the pathogenesis modulating the expression of oncogenes and/or tumor suppressor genes and interacting with SIRT1, which crucially impacts the circadian clock circuitry. Hence, we aimed to appraise if and how macroH2A1 regulated the expression patterns of circadian genes in the setting of NAFLD-associated HCC. We took advantage of an in vitro model of liver cancer represented by HepG2 (human hepatocarcinoma) cells stably knocked down for macroH2A1 and conducted whole transcriptome profiling and deep phenotyping analysis. We found up-regulation of PER1 along with several deregulated circadian genes, enriching several important pathways and functions related to cancer onset and progression, such as epithelial-to-mesenchymal transition, cell cycle deregulation, and DNA damage. PER1 silencing partially mitigated the malignant phenotype induced by the loss of macroH2A1 in HCC cells. In conclusion, our findings suggest a modulatory role for the core circadian protein PER1 in liver carcinogenesis in the context of a lack of the macroH2A1 epigenetic and transcriptional landscape. Keywords: circadian; cancer; biological clock; HCC; PER1 Author: Annalucia Carbone, Elisabetta De Santis, Olga Cela, Vincenzo Giambra, Luca Miele, Giuseppe Marrone, Antonio Grieco, Marcus Buschbeck, Nazzareno Capitanio, Tommaso Mazza and Gianluigi Mazzoccoli Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Tue Aug 24 10:50:05 2021 CEST ModDate: Tue Aug 24 10:53:20 2021 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 21 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 3506154 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- EXVEHZ+VnURWPalladioL Type 1 Custom yes yes no 10 0 WZLPTR+URWPalladioL-Ital Type 1 Custom yes yes no 15 0 SUPIFP+URWPalladioL-Bold Type 1 Custom yes yes no 20 0 DKLMDS+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 EBFBTV+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 32 0 MCBCCI+SourceSansRoman-Regular Type 1C Custom yes yes no 54 0 MCBCCM+SourceSansRoman.613wght Type 1C Custom yes yes no 58 0 MCBCCO+SourceSansRoman-Bold Type 1C Custom yes yes no 62 0 KESBIZ+EURM10 Type 1 Builtin yes yes no 74 0 IFEDSM+CMSY10 Type 1 Builtin yes yes no 81 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 104 0 KFNDPP+PalatinoLinotype CID TrueType Identity-H yes yes yes 107 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 113 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 116 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 136 0 KFNDPP+PalatinoLinotype CID TrueType Identity-H yes yes yes 139 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 145 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 148 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 162 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 165 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 168 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 183 0 KFNDPP+PalatinoLinotype CID TrueType Identity-H yes yes yes 186 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 192 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 195 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 209 0 KFNDPP+PalatinoLinotype CID TrueType Identity-H yes yes yes 212 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 218 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 221 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 248 0 KFNDPP+PalatinoLinotype CID TrueType Identity-H yes yes yes 251 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 257 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 260 0 KFNDPO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 282 0 KFNEAA+PalatinoLinotype TrueType WinAnsi yes yes no 285 0 KFNEBB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 288 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-03-11 02:35:27 CET RepresentationInformation: 270598.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2023-03-10 17:11:06 CET Size: 3506154 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 630 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@44c8afef Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7b69c6ba Children: Item: Title: Primary Datasets Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@46daef40 Item: Title: Cell Culture Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@12f41634 Item: Title: Generation of Lentiviral Vectors Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@13c27452 Item: Title: RNA-Seq Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Item: Title: Analysis for Functional Categorization of the Most Significant Pathways Enriched by Deregulated Circadian Genes Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Item: Title: Knockdown of PER1 by RNA Interference Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: Western Blotting Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: Cell Proliferation Assay Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: Cell Migration Assay Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Item: Title: Flow Cytometry Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Children: Item: Title: Cell Cycle Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Item: Title: Half Maximal Inhibitory Concentration (IC50) of Cisplatin Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@a38d7a3 Item: Title: DNA Damage and Apoptosis Assay with and without Cisplatin Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77f99a05 Item: Title: Mitochondria and Cellular Respiration Measurements Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@63440df3 Item: Title: Statistical Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@3aeaafa6 Item: Title: Results Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@76a3e297 Children: Item: Title: Loss of MacroH2A1 Histone Variants Changes the Expression of Circadian Genes in HCC Cells Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@4d3167f4 Item: Title: Circadian Genes Deregulated upon MacroH2A1 Knockout Enrich Important Pathways in HCC Cells Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@ed9d034 Item: Title: MacroH2A1 Histone Knockdown in HCC Alters the Expression of Proteins Entailed in Key Cell Functions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@6121c9d6 Item: Title: Knockdown of MacroH2A1 Histone Modulates Proliferative and Migratory Capacity of HCC Cells Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@87f383f Item: Title: MacroH2A1 Histone Loss Impacts Cell Cycle, Anticancer Drug Sensitivity, DNA Damage/Apoptosis in HCC Cells Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@4eb7f003 Item: Title: Loss of MacroH2A1 Decreases Mitochondrial Oxidative Phosphorylation Efficiency Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@eafc191 Item: Title: Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@612fc6eb Item: Title: Conclusions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1060b431 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@612679d6 Info: Title: The Histone Variant MacroH2A1 Impacts Circadian Gene Expression and Cell Phenotype in an In Vitro Model of Hepatocellular Carcinoma Author: Annalucia Carbone, Elisabetta De Santis, Olga Cela, Vincenzo Giambra, Luca Miele, Giuseppe Marrone, Antonio Grieco, Marcus Buschbeck, Nazzareno Capitanio, Tommaso Mazza and Gianluigi Mazzoccoli Subject: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. A foremost risk factor for HCC is obesity/metabolic syndrome-related non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH), which is prompted by remarkable changes in transcription patterns of genes enriching metabolic, immune/inflammatory, and circadian pathways. Epigenetic mechanisms play a role in NAFLD-associated HCC, and macroH2A1, a variant of histone H2A, is involved in the pathogenesis modulating the expression of oncogenes and/or tumor suppressor genes and interacting with SIRT1, which crucially impacts the circadian clock circuitry. Hence, we aimed to appraise if and how macroH2A1 regulated the expression patterns of circadian genes in the setting of NAFLD-associated HCC. We took advantage of an in vitro model of liver cancer represented by HepG2 (human hepatocarcinoma) cells stably knocked down for macroH2A1 and conducted whole transcriptome profiling and deep phenotyping analysis. We found up-regulation of PER1 along with several deregulated circadian genes, enriching several important pathways and functions related to cancer onset and progression, such as epithelial-to-mesenchymal transition, cell cycle deregulation, and DNA damage. PER1 silencing partially mitigated the malignant phenotype induced by the loss of macroH2A1 in HCC cells. In conclusion, our findings suggest a modulatory role for the core circadian protein PER1 in liver carcinogenesis in the context of a lack of the macroH2A1 epigenetic and transcriptional landscape. 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