1 1176 180 CONTROL OF APOBEC3B INDUCTION AND CCCDNA DECAY BY NF-KAPPAB AND MIR-138-5P. BACKGROUND & AIMS: IMMUNE-MEDIATED INDUCTION OF CYTIDINE DEAMINASE APOBEC3B (A3B) EXPRESSION LEADS TO HBV COVALENTLY CLOSED CIRCULAR DNA (CCCDNA) DECAY. HERE, WE AIMED TO DECIPHER THE SIGNALLING PATHWAY(S) AND REGULATORY MECHANISM(S) INVOLVED IN A3B INDUCTION AND RELATED HBV CONTROL. METHODS: DIFFERENTIATED HEPARG CELLS (DHEPARG) KNOCKED-DOWN FOR NF-KAPPAB SIGNALLING COMPONENTS, TRANSFECTED WITH SIRNA OR MICRO RNAS (MIRNA), AND PRIMARY HUMAN HEPATOCYTES +/- HBV OR HBVDELTAX OR HBV-RFP, WERE TREATED WITH LYMPHOTOXIN BETA RECEPTOR (LTBETAR)-AGONIST (BS1). THE BIOLOGICAL OUTCOMES WERE ANALYSED BY REVERSE TRANSCRIPTASE-QPCR, IMMUNOBLOTTING, LUCIFERASE ACTIVITY, CHROMATIN IMMUNE PRECIPITATION, ELECTROPHORETIC MOBILITY-SHIFT ASSAY, TARGETED-BISULFITE-, MIRNA-, RNA-, GENOME-SEQUENCING, AND MASS-SPECTROMETRY. RESULTS: WE FOUND THAT CANONICAL AND NON-CANONICAL NF-KAPPAB SIGNALLING PATHWAYS ARE MANDATORY FOR A3B INDUCTION AND ANTI-HBV EFFECTS. THE DEGREE OF IMMUNE-MEDIATED A3B PRODUCTION IS INDEPENDENT OF A3B PROMOTER DEMETHYLATION BUT IS CONTROLLED POST-TRANSCRIPTIONALLY BY THE MIRNA 138-5P EXPRESSION (HSA-MIR-138-5P), PROMOTING A3B MRNA DECAY. HSA-MIR-138-5P OVER-EXPRESSION REDUCED A3B LEVELS AND ITS ANTIVIRAL EFFECTS. OF NOTE, ESTABLISHED INFECTION INHIBITED BS1-INDUCED A3B EXPRESSION THROUGH EPIGENETIC MODULATION OF A3B PROMOTER. TWELVE DAYS OF TREATMENT WITH A LTBETAR-SPECIFIC AGONIST BS1 IS SUFFICIENT TO REDUCE THE CCCDNA POOL BY 80% WITHOUT INDUCING SIGNIFICANT DAMAGES TO A SUBSET OF CANCER-RELATED HOST GENES. INTERESTINGLY, THE A3B-MEDIATED EFFECT ON HBV IS INDEPENDENT OF THE TRANSCRIPTIONAL ACTIVITY OF CCCDNA AS WELL AS ON RCDNA SYNTHESIS. CONCLUSIONS: ALTOGETHER, A3B REPRESENTS THE ONLY DESCRIBED ENZYME TO TARGET BOTH TRANSCRIPTIONALLY ACTIVE AND INACTIVE CCCDNA. THUS, INHIBITING HSA-MIR-138-5P EXPRESSION SHOULD BE CONSIDERED IN THE COMBINATORIAL DESIGN OF NEW THERAPIES AGAINST HBV, ESPECIALLY IN THE CONTEXT OF IMMUNE-MEDIATED A3B INDUCTION. LAY SUMMARY: IMMUNE-MEDIATED INDUCTION OF CYTIDINE DEAMINASE APOBEC3B IS TRANSCRIPTIONALLY REGULATED BY NF-KAPPAB SIGNALLING AND POST-TRANSCRIPTIONALLY DOWNREGULATED BY HSA-MIR-138-5P EXPRESSION, LEADING TO CCCDNA DECAY. TIMELY CONTROLLED APOBEC3B-MEDIATED CCCDNA DECAY OCCURS INDEPENDENTLY OF CCCDNA TRANSCRIPTIONAL ACTIVITY AND WITHOUT DAMAGE TO A SUBSET OF CANCER-RELATED GENES. THUS, APOBEC3B-MEDIATED CCCDNA DECAY COULD OFFER AN EFFICIENT THERAPEUTIC ALTERNATIVE TO TARGET HEPATITIS B VIRUS CHRONIC INFECTION.	2021	
                                                                                                                                                                                                                                                                                        
2 5678  47 SHORT HAIRPIN RNA INDUCES METHYLATION OF HEPATITIS B VIRUS COVALENTLY CLOSED CIRCULAR DNA IN HUMAN HEPATOMA CELLS. SMALL INTERFERING RNAS NOT ONLY MODULATE GENE EXPRESSION AT A POST-TRANSCRIPTIONAL LEVEL, BUT ALSO INDUCE TRANSCRIPTIONAL GENE SILENCING BY RNA INTERFERENCE-MEDIATED HETEROCHROMATIN FORMATION AND RNA-DIRECTED DNA METHYLATION (RDDM). HOWEVER, ALTHOUGH ESTABLISHED IN PLANTS, THERE HAVE BEEN CONTROVERSIES WHETHER RDDM OPERATES IN MAMMALS. HEPATITIS B VIRUS (HBV) COVALENTLY CLOSED CIRCULAR DNA (CCCDNA) SERVES AS A TEMPLATE FOR VIRAL RNA TRANSCRIPTION, AND TRANSCRIPTIONAL ACTIVITY OF HBV CCCDNA IS REGULATED BY METHYLATION IN PATIENTS WITH CHRONIC HBV INFECTION. IN THIS STUDY, WE STABLY EXPRESSED SHORT HAIRPIN RNA (SHRNA) AGAINST HBV IN HUMAN HEPATOMA CELLS TO DETERMINE WHETHER SHRNA INDUCES METHYLATION OF HBV CCCDNA. HEPAD38 CELLS WHICH PERMIT REPLICATION OF HBV UNDER CONTROL OF TETRACYCLINE-RESPONSIVE PROMOTER WERE TRANSDUCED WITH LENTIVIRAL VECTORS WHICH ENCODE SH-1580, A SHRNA AGAINST THE HEPATITIS B VIRAL PROTEIN HBX. BISULFITE SEQUENCING PCR ANALYSIS REVEALED THAT SH-1580 INDUCED CPG METHYLATIONS AT A HIGHER RATE COMPARED TO CONTROL (31.3% VS. 12.8%, P<0.05). THE SH-1580-INDUCED CPG METHYLATION WAS LOCALIZED NEAR THE TARGET SEQUENCE OF SH-1580 IN MORE THAN A HALF OF THE CLONES. METHYLATION-INDUCED TRANSCRIPTIONAL SUPPRESSION WAS CONFIRMED BY IN VITRO TRANSCRIPTION ASSAY. THESE RESULTS CONFIRM THE FEASIBILITY OF RDDM OF HBV CCCDNA IN HUMAN CELLS. LENTIVIRAL VECTOR-MEDIATED TRANSFER OF SHRNA MAY BE USED AS A TOOL FOR NOVEL TRANSCRIPTIONAL MODULATION BY EPIGENETIC MODIFICATION OF HBV CCCDNA.	2013	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
3 2837  52 FORKHEAD O TRANSCRIPTION FACTOR 4 RESTRICTS HBV COVALENTLY CLOSED CIRCULAR DNA TRANSCRIPTION AND HBV REPLICATION THROUGH GENETIC DOWNREGULATION OF HEPATOCYTE NUCLEAR FACTOR 4 ALPHA AND EPIGENETIC SUPPRESSION OF COVALENTLY CLOSED CIRCULAR DNA VIA INTERACTING WITH PROMYELOCYTIC LEUKEMIA PROTEIN. NUCLEAR LOCATED HEPATITIS B VIRUS (HBV) COVALENTLY CLOSED CIRCULAR DNA (CCCDNA) REMAINS THE KEY OBSTACLE TO CURE CHRONIC HEPATITIS B (CHB). IN OUR PREVIOUS INVESTIGATION, IT WAS FOUND THAT FOXO4 COULD INHIBIT HBV CORE PROMOTER ACTIVITY THROUGH DOWNREGULATING THE EXPRESSION OF HNF4ALPHA. HOWEVER, THE EXACT MECHANISMS WHEREBY FOXO4 INHIBITS HBV REPLICATION, ESPECIALLY ITS EFFECT ON CCCDNA, REMAIN UNCLEAR. HERE, OUR DATA FURTHER REVEALED THAT FOXO4 COULD EFFECTIVELY INHIBIT CCCDNA MEDIATED TRANSCRIPTION AND HBV REPLICATION WITHOUT AFFECTING CCCDNA LEVEL. MECHANISTIC STUDY SHOWED THAT FOXO4 COULD CAUSE EPIGENETIC SUPPRESSION OF CCCDNA. ALTHOUGH FOXO4-MEDIATED DOWNREGULATION OF HNF4ALPHA CONTRIBUTED TO INHIBITING HBV CORE PROMOTER ACTIVITY, IT HAD LITTLE EFFECT ON CCCDNA EPIGENETIC REGULATION. FURTHER, IT WAS FOUND THAT FOXO4 COULD COLOCALIZE WITHIN PROMYELOCYTIC LEUKEMIA PROTEIN (PML) NUCLEAR BODIES AND INTERACT WITH PML. OF NOTE, PML WAS REVEALED TO BE CRITICAL FOR FOXO4-MEDIATED INHIBITION OF CCCDNA EPIGENETIC MODIFICATION AND OF THE FOLLOWING CCCDNA TRANSCRIPTION AND HBV REPLICATION. FURTHERMORE, FOXO4 WAS FOUND TO BE DOWNREGULATED IN HBV-INFECTED HEPATOCYTES AND HUMAN LIVER TISSUES, AND IT WAS NEGATIVELY CORRELATED WITH CCCDNA TRANSCRIPTIONAL ACTIVITY IN CHB PATIENTS. TOGETHER, THESE FINDINGS HIGHLIGHT THE ROLE OF FOXO4 IN SUPPRESSING CCCDNA TRANSCRIPTION AND HBV REPLICATION VIA GENETIC DOWNREGULATION OF HNF4ALPHA AND EPIGENETIC SUPPRESSION OF CCCDNA THROUGH INTERACTING WITH PML. TARGETING FOXO4 MAY PRESENT AS A NEW THERAPEUTIC STRATEGY AGAINST CHRONIC HBV INFECTION. IMPORTANCE HBV CCCDNA IS A DETERMINING FACTOR FOR VIRAL PERSISTENCE AND THE MAIN OBSTACLE FOR A CURE OF CHRONIC HEPATITIS B. STRATEGIES THAT TARGET CCCDNA DIRECTLY ARE THEREFORE OF GREAT IMPORTANCE IN CONTROLLING PERSISTENT HBV INFECTION. IN PRESENT INVESTIGATION, WE FOUND THAT FOXO4 COULD EFFICIENTLY SUPPRESS CCCDNA TRANSCRIPTION AND HBV REPLICATION WITHOUT AFFECTING THE LEVEL OF CCCDNA ITSELF. FURTHER, OUR DATA REVEALED THAT FOXO4 MIGHT INHIBIT CCCDNA FUNCTION VIA A TWO-PART MECHANISM: ONE IS TO EPIGENETICALLY SUPPRESS CCCDNA TRANSCRIPTION VIA INTERACTING WITH PML, AND THE OTHER IS TO INHIBIT HBV CORE PROMOTER ACTIVITY VIA THE GENETIC DOWNREGULATION OF HNF4ALPHA. OF NOTE, HBV MIGHT DAMPEN THE EXPRESSION OF FOXO4 FOR ITS OWN PERSISTENT INFECTION. WE PROPOSE THAT MANIPULATION OF FOXO4 MAY PRESENT AS A POTENTIAL THERAPEUTIC STRATEGY AGAINST CHRONIC HBV INFECTION.	2022	
                                                                                   
4 4771  46 NUCLEAR SENSOR INTERFERON-INDUCIBLE PROTEIN 16 INHIBITS THE FUNCTION OF HEPATITIS B VIRUS COVALENTLY CLOSED CIRCULAR DNA BY INTEGRATING INNATE IMMUNE ACTIVATION AND EPIGENETIC SUPPRESSION. BACKGROUND AND AIMS: NUCLEAR-LOCATED COVALENTLY CLOSED CIRCULAR DNA (CCCDNA) OF HEPATITIS B VIRUS (HBV) IS A DETERMINING FACTOR FOR HBV PERSISTENCE AND THE KEY OBSTACLE FOR A CURE OF CHRONIC HEPATITIS B. HOWEVER, IT REMAINS UNCLEAR WHETHER AND HOW THE HOST IMMUNE SYSTEM SENSES HBV CCCDNA AND ITS BIOLOGICAL CONSEQUENCES. APPROACH AND RESULTS: HERE, WE DEMONSTRATED THAT INTERFERON-INDUCIBLE PROTEIN 16 (IFI16) COULD SERVE AS A UNIQUE INNATE SENSOR TO RECOGNIZE AND BIND TO HBV CCCDNA IN HEPATIC NUCLEI, LEADING TO THE INHIBITION OF CCCDNA TRANSCRIPTION AND HBV REPLICATION. MECHANISTICALLY, OUR DATA SHOWED THAT IFI16 PROMOTED THE EPIGENETIC SUPPRESSION OF HBV CCCDNA BY TARGETING AN INTERFERON-STIMULATED RESPONSE ELEMENT (ISRE) PRESENT IN CCCDNA. IT IS OF INTEREST THAT THIS ISRE WAS ALSO REVEALED TO PLAY AN IMPORTANT ROLE IN IFI16-ACTIVATED TYPE I INTERFERON RESPONSES. FURTHERMORE, OUR DATA REVEALED THAT HBV COULD DOWN-REGULATE THE EXPRESSION LEVEL OF IFI16 IN HEPATOCYTES, AND THERE WAS A NEGATIVE CORRELATION BETWEEN IFI16 AND HBV TRANSCRIPTS IN LIVER BIOPSIES, SUGGESTING THE POSSIBLE ROLE OF IFI16 IN SUPPRESSING CCCDNA FUNCTION UNDER PHYSIOLOGICAL CONDITIONS. CONCLUSIONS: THE NUCLEAR SENSOR IFI16 SUPPRESSES CCCDNA FUNCTION BY INTEGRATING INNATE IMMUNE ACTIVATION AND EPIGENETIC REGULATION BY TARGETING THE ISRE OF CCCDNA, AND IFI16 MAY PRESENT AS A THERAPEUTIC TARGET AGAINST HBV INFECTION.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     
5 6448  54 THERAPEUTIC SHUTDOWN OF HBV TRANSCRIPTS PROMOTES REAPPEARANCE OF THE SMC5/6 COMPLEX AND SILENCING OF THE VIRAL GENOME IN VIVO. OBJECTIVE: THERAPEUTIC STRATEGIES SILENCING AND REDUCING THE HEPATITIS B VIRUS (HBV) RESERVOIR, THE COVALENTLY CLOSED CIRCULAR DNA (CCCDNA), HAVE THE POTENTIAL TO CURE CHRONIC HBV INFECTION. WE AIMED TO INVESTIGATE THE IMPACT OF SMALL INTERFERRING RNA (SIRNA) TARGETING ALL HBV TRANSCRIPTS OR PEGYLATED INTERFERON-ALPHA (PEG-IFNALPHA) ON THE VIRAL REGULATORY HBX PROTEIN AND THE STRUCTURAL MAINTENANCE OF CHROMOSOME 5/6 COMPLEX (SMC5/6), A HOST FACTOR SUPPRESSING CCCDNA TRANSCRIPTION. IN PARTICULAR, WE ASSESSED WHETHER INTERVENTIONS LOWERING HBV TRANSCRIPTS CAN ACHIEVE AND MAINTAIN SILENCING OF CCCDNA TRANSCRIPTION IN VIVO. DESIGN: HBV-INFECTED HUMAN LIVER CHIMERIC MICE WERE TREATED WITH SIRNA OR PEG-IFNALPHA. VIROLOGICAL AND HOST CHANGES WERE ANALYSED AT THE END OF TREATMENT AND DURING THE REBOUND PHASE BY QUALITATIVE PCR, ELISA, IMMUNOBLOTTING AND CHROMATIN IMMUNOPRECIPITATION. RNA IN SITU HYBRIDISATION WAS COMBINED WITH IMMUNOFLUORESCENCE TO DETECT SMC6 AND HBV RNAS AT SINGLE CELL LEVEL. THE ENTRY INHIBITOR MYRCLUDEX-B WAS USED DURING THE REBOUND PHASE TO AVOID NEW INFECTION EVENTS. RESULTS: BOTH SIRNA AND PEG-IFNALPHA STRONGLY REDUCED ALL HBV MARKERS, INCLUDING HBX LEVELS, THUS ENABLING THE REAPPEARANCE OF SMC5/6 IN HEPATOCYTES THAT ACHIEVED HBV-RNA NEGATIVISATION AND SMC5/6 ASSOCIATION WITH THE CCCDNA. ONLY IFN REDUCED CCCDNA LOADS AND ENHANCED IFN-STIMULATED GENES. HOWEVER, THE ANTIVIRAL EFFECTS DID NOT PERSIST OFF TREATMENT AND SMC5/6 WAS AGAIN DEGRADED. REMARKABLY, THE BLOCKADE OF VIRAL ENTRY THAT STARTED AT THE END OF TREATMENT HINDERED RENEWED DEGRADATION OF SMC5/6. CONCLUSION: THESE RESULTS REVEAL THAT THERAPEUTICS ABROGATING ALL HBV TRANSCRIPTS INCLUDING HBX PROMOTE EPIGENETIC SUPPRESSION OF THE HBV MINICHROMOSOME, WHEREAS STRATEGIES PROTECTING THE HUMAN HEPATOCYTES FROM REINFECTION ARE NEEDED TO MAINTAIN CCCDNA SILENCING.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         
6 3728  45 INHIBITION OF REPLICATION OF HEPATITIS B VIRUS USING TRANSCRIPTIONAL REPRESSORS THAT TARGET THE VIRAL DNA. BACKGROUND: CHRONIC INFECTION WITH HEPATITIS B VIRUS (HBV) IS A SERIOUS GLOBAL HEALTH PROBLEM. PERSISTENCE OF THE VIRUS OCCURS AS A RESULT OF STABILITY OF THE REPLICATION INTERMEDIATE COMPRISING COVALENTLY CLOSED CIRCULAR DNA (CCCDNA). DEVELOPMENT OF DRUGS THAT ARE CAPABLE OF DISABLING THIS CCCDNA IS VITAL. METHODS: TO INVESTIGATE AN EPIGENETIC APPROACH TO INACTIVATING VIRAL DNA, WE ENGINEERED TRANSCRIPTIONAL REPRESSORS THAT COMPRISE AN HBV DNA-BINDING DOMAIN OF TRANSCRIPTION ACTIVATOR LIKE EFFECTORS (TALES) AND A FUSED KRUPPEL ASSOCIATED BOX (KRAB). THESE REPRESSOR TALES (RTALES) TARGETED THE VIRAL SURFACE OPEN READING FRAME AND WERE PLACED UNDER TRANSCRIPTION CONTROL OF CONSTITUTIVELY ACTIVE OR LIVER-SPECIFIC PROMOTERS. RESULTS: EVALUATION IN CULTURED CELLS AND FOLLOWING HYDRODYNAMIC INJECTION OF MICE REVEALED THAT THE RTALES SIGNIFICANTLY INHIBITED PRODUCTION OF MARKERS OF HBV REPLICATION WITHOUT EVIDENCE OF HEPATOTOXICITY. INCREASED METHYLATION OF HBV DNA AT CPG ISLAND II SHOWED THAT THE RTALES CAUSED INTENDED EPIGENETIC MODIFICATION. CONCLUSIONS: EPIGENETIC MODIFICATION OF HBV DNA IS A NEW AND EFFECTIVE MEANS OF INACTIVATING THE VIRUS IN VIVO. THE APPROACH HAS THERAPEUTIC POTENTIAL AND AVOIDS POTENTIALLY PROBLEMATIC UNINTENDED MUTAGENESIS OF GENE EDITING.	2019	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   
7 3128  41 GIPC-REGULATED IGFBP-3 PROMOTES HSC MIGRATION IN VITRO AND PORTAL HYPERTENSION IN VIVO THROUGH A BETA1-INTEGRIN PATHWAY. BACKGROUND & AIMS: TRANSFORMING GROWTH FACTOR (TGF-BETA)-INDUCED ACTIVATION OF QUIESCENT HEPATIC STELLATE CELLS (HSCS) AND THEIR TRANSFORMATION TO MYOFIBROBLASTS IS A KEY EVENT IN LIVER FIBROSIS AND PORTAL HYPERTENSION. GIPC (ALSO REFERRED TO AS SYNECTIN) IS A DOWNSTREAM SIGNAL ACTIVATION MOLECULE OF TGF-BETA AND OTHER RECEPTORS. IN THIS STUDY, WE SOUGHT TO IDENTIFY NOVEL GENES TARGETED BY TGF-BETA AND GIPC AND ELUCIDATE IF AND HOW THEY MAY CONTRIBUTE TO LIVER FIBROSIS. METHODS: WE PERFORMED SEQUENTIAL MESSENGER RNA SEQUENCING ANALYSIS ON TGF-BETA-STIMULATED HSCS AND THEN ON TGF-BETA-STIMULATED HSCS IN THE PRESENCE AND ABSENCE OF GIPC ALSO REFERRED TO AS SYNECTIN (GIPC) KNOCKDOWN. INSULIN-LIKE GROWTH FACTOR BINDING PROTEIN-3 (IGFBP-3) TRANSPORT PROTEIN EMERGED AS A TOP ACTIVATION TARGET OF BOTH TGF-BETA AND GIPC. QUANTITATIVE POLYMERASE CHAIN REACTION, ENZYME-LINKED IMMUNOSORBENT ASSAY, TARGETED CHROMATIN IMMUNOPRECIPITATION, AND WESTERN BLOT ANALYSIS WERE DONE FOR FURTHER CONFIRMATION. RESULTS: IGFBP-3, AN INSULIN GROWTH FACTOR TRANSPORT PROTEIN, EMERGED AS A TOP ACTIVATION TARGET OF BOTH TGF-BETA AND GIPC, WHICH WAS CONFIRMED BY QUANTITATIVE POLYMERASE CHAIN REACTION, ENZYME-LINKED IMMUNOSORBENT ASSAY, AND WESTERN BLOT ANALYSIS. TARGETED CHROMATIN IMMUNOPRECIPITATION SHOWED THAT GIPC INCREASES THE HISTONE 3 LYSINE 27 (H3K27) ACETYLATION ACTIVATING MARK AND CONCURRENTLY DECREASES THE H3K27 INHIBITORY TRIMETHYLATION (H3K27M3) MARK, PROVIDING AN EPIGENETIC CORRELATE TO THE GENE REGULATION CHANGES. IN VIVO, GLOBAL KNOCKOUT OF IGFBP-3 MICE RESULTED IN ATTENUATION OF HSC ACTIVATION MARKERS AND ATTENUATION OF PORTAL PRESSURE IN RESPONSE TO CHRONIC LIVER INJURY MODELS. ANALYSIS OF SERUM LEVELS FROM CIRRHOTIC PATIENTS ALSO SHOWED AN IGFBP-3 INCREASE OF MORE THAN 2-FOLD COMPARED WITH HEALTHY CONTROLS. FINALLY, IN VITRO MECHANISM STUDIES SHOWED THAT IGFBP-3 PROMOTES HSC MIGRATION THROUGH INTEGRIN-DEPENDENT PHOSPHORYLATION OF PROTEIN KINASE B. CONCLUSIONS: TGF-BETA UP-REGULATES IGFBP-3 THROUGH GIPC, LEADING TO INCREASED HSC MIGRATION IN VITRO AND PROMOTES PORTAL HYPERTENSION IN VIVO. THESE STUDIES SUPPORT THE ROLE OF IGFBP-3 AS A POTENTIAL PATHOPHYSIOLOGIC TARGET OR BIOMARKER IN CHRONIC LIVER DISEASE.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
8 3252  51 HEPATITIS B VIRUS SUPPRESSES COMPLEMENT C9 SYNTHESIS BY LIMITING THE AVAILABILITY OF TRANSCRIPTION FACTOR USF-1 AND INHIBITS FORMATION OF MEMBRANE ATTACK COMPLEX: IMPLICATIONS IN DISEASE PATHOGENESIS. BACKGROUND: THE COMPLEMENT SYSTEM FUNCTIONS PRIMARILY AS A FIRST-LINE HOST DEFENSE AGAINST INVADING MICROBES, INCLUDING VIRUSES. HOWEVER, THE INTERACTION OF HEPATITIS B VIRUS (HBV) WITH THE COMPLEMENT-COMPONENTS DURING CHRONIC HBV INFECTION REMAINS LARGELY UNKNOWN. WE INVESTIGATED THE MECHANISM BY WHICH HBV INHIBITS THE FORMATION OF CYTOLYTIC COMPLEMENT MEMBRANE-ATTACK COMPLEX (MAC) AND STUDIED ITS IMPACT ON MAC-MEDIATED MICROBICIDAL ACTIVITY AND DISEASE PATHOGENESIS. METHODS: BLOOD/LIVER TISSUES WERE COLLECTED FROM CHRONICALLY HBV-INFECTED PATIENTS AND CONTROLS. HEPG2(HNTCP) CELLS WERE INFECTED WITH HBV PARTICLES AND HUH7 CELLS WERE TRANSFECTED WITH FULL-LENGTH LINEAR HBV-MONOMER OR PLASMIDS CONTAINING DIFFERENT HBV-ORFS AND EXPRESSION OF COMPLEMENT COMPONENTS OR OTHER HOST GENES WERE EVALUATED. ADDITIONALLY, ELISA, REAL-TIME PCR, WESTERN BLOT, BIOINFORMATICS ANALYSIS, GENE OVEREXPRESSION/KNOCK-DOWN, MUTAGENESIS, CHROMATIN IMMUNOPRECIPITATION, EPIGENETIC STUDIES, IMMUNOFLUORESCENCE, AND QUANTIFICATION OF SERUM HBV-DNA, BACTERIAL-DNA AND ENDOTOXIN WERE PERFORMED. RESULTS: AMONG THE MAC COMPONENTS (C5B-C9), SIGNIFICANT REDUCTION WAS NOTED IN THE EXPRESSION OF C9, THE MAJOR CONSTITUENT OF MAC, IN HBV-INFECTED HEPG2(HNTCP) CELLS AND IN HUH7 CELLS TRANSFECTED WITH FULL-LENGTH HBV AS WELL AS HBX. C9 LEVEL WAS ALSO MARKED LOW IN SERA/LIVER OF CHRONIC HEPATITIS B (CHB) AND IMMUNE-TOLERANT (IT) PATIENTS THAN INACTIVE CARRIERS AND HEALTHY CONTROLS. HBX STRONGLY REPRESSED C9-PROMOTER ACTIVITY IN HUH7 CELLS BUT CPG-ISLAND WAS NOT DETECTED IN C9-PROMOTER. WE IDENTIFIED USF-1 AS THE KEY TRANSCRIPTION FACTOR THAT DRIVES C9 EXPRESSION AND DEMONSTRATED THAT HBX-INDUCED HYPERMETHYLATION OF USF-1-PROMOTER IS THE LEADING CAUSE OF USF-1 DOWNREGULATION THAT IN TURN DIMINISHED C9 TRANSCRIPTION. REDUCED MAC FORMATION AND IMPAIRED LYSIS OF HBV-TRANSFECTED HUH7 AND BACTERIAL CELLS WERE OBSERVED FOLLOWING INCUBATION OF THESE CELLS WITH C9-DEFICIENT CHB SERA BUT WAS REVERSED UPON C9 SUPPLEMENTATION. SIGNIFICANT INVERSE CORRELATION WAS NOTED BETWEEN C9 CONCENTRATION AND HBV-DNA, BACTERIAL-DNA AND ENDOTOXIN CONTENT IN HBV-INFECTED PATIENTS. ONE-YEAR TENOFOVIR THERAPY RESULTED IN IMPROVEMENT IN C9 LEVEL AND DECLINE IN VIRAL/BACTERIAL/ENDOTOXIN LOAD IN CHB PATIENTS. CONCLUSION: COLLECTIVELY, HBX SUPPRESSED C9 TRANSCRIPTION BY RESTRICTING THE AVAILABILITY OF USF-1 THROUGH HYPERMETHYLATION OF USF-1-PROMOTER AND CONSEQUENTLY HINDER THE FORMATION AND LYTIC FUNCTIONS OF MAC. EARLY THERAPY IS NEEDED FOR BOTH CHB AND IT TO NORMALIZE THE ABERRANT COMPLEMENT PROFILE AND CONTAIN VIRAL AND BACTERIAL INFECTION AND LIMIT DISEASE PROGRESSION.	2022	

9 3249  38 HEPATITIS B VIRUS HIJACKS CTHRC1 TO EVADE HOST IMMUNITY AND MAINTAIN REPLICATION. HEPATITIS B VIRUS (HBV) INFECTION CAUSES ACUTE AND CHRONIC LIVER DISEASES, BUT IS NOT DIRECTLY CYTOPATHIC. LIVER INJURY RESULTS FROM REPEATED ATTEMPTS OF THE CELLULAR IMMUNE RESPONSE SYSTEM TO CONTROL THE VIRAL INFECTION. HERE, WE INVESTIGATE THE ROLES OF CELLULAR FACTORS AND SIGNALING PATHWAYS INVOLVED IN THE REGULATION OF HBV REPLICATION TO REVEAL THE MECHANISM UNDERLYING HBV INFECTION AND PATHOGENESIS. WE SHOW THAT COLLAGEN TRIPLE HELIX REPEAT CONTAINING 1 (CTHRC1) EXPRESSION IS ELEVATED IN HBV-INFECTED PATIENTS AND IN HBV-TRANSFECTED CELLS THROUGH EPIGENETIC MODIFICATION AND TRANSCRIPTIONAL REGULATION. CTHRC1 FACILITATES HBV REPLICATION IN CULTURED CELLS AND BALB/C MICE BY ACTIVATING THE PKCALPHA/ERK/JNK/C-JUN CASCADE TO REPRESS THE IFN/JAK/STAT PATHWAY. HBV-ACTIVATED CTHRC1 DOWNREGULATES THE ACTIVITY OF TYPE I INTERFERON (IFN), THE PRODUCTION OF IFN-STIMULATED GENES (ISGS), AND THE PHOSPHORYLATION OF SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION 1/2 (STAT1/2), WHEREAS IT UPREGULATES THE PHOSPHORYLATION AND UBIQUITINATION OF TYPE I IFN RECEPTORS (IFNARALPHA/BETA). THUS, OUR RESULTS SHOW THAT HBV USES A NOVEL MECHANISM TO HIJACK CELLULAR FACTORS AND SIGNAL CASCADES IN ORDER TO EVADE HOST ANTIVIRAL IMMUNITY AND MAINTAIN PERSISTENT INFECTION. WE ALSO DEMONSTRATE THAT CTHRC1 HAS A NOVEL ROLE IN VIRAL INFECTION.	2015	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     
10 2432  38 EPIGENETIC SILENCING OF MIR-708 ENHANCES NF-KAPPAB SIGNALING IN CHRONIC LYMPHOCYTIC LEUKEMIA. MICRORNAS (MIRNAS) ARE POST-TRANSCRIPTIONAL REGULATORS OF GENE EXPRESSION AND THEIR DEREGULATION IS INVOLVED IN TUMOR DEVELOPMENT. EPIGENETIC GENE SILENCING IN CANCER BY DNA METHYLATION CONTRIBUTES TO THE SILENCING OF TUMOR-SUPPRESSOR GENES, INCLUDING MIRNAS. WE HAVE RECENTLY SHOWN THAT THE PROMOTER OF MIR-708 IS ABERRANTLY METHYLATED IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). TO CHARACTERIZE THE MOLECULAR SIGNALING NETWORKS THAT ARE INFLUENCED BY MIR-708, WE PERFORMED A LUCIFERASE-BASED SCREEN EVALUATING THE EFFECTS OF ECTOPIC MIR-708 EXPRESSION ON LEUKEMIA-RELEVANT SIGNALING PATHWAYS. WE FOUND THAT MIR-708 STRONGLY REPRESSED NF-KAPPAB SIGNALING, A PATHWAY KNOWN TO BE DEREGULATED IN CLL. AMONG THE PREDICTED MIR-708 TARGETS WAS IKKBETA (INHIBITOR OF KAPPA LIGHT POLYPEPTIDE GENE ENHANCER IN B CELLS, KINASE-BETA/IKBKB), A KEY KINASE FACILITATING NF-KAPPAB SIGNALING. WE VALIDATED THE INTERACTION OF MIR-708 WITH THE 3'-UNTRANSLATED REGION OF IKKBETA AND FOUND THAT MIR-708 OVEREXPRESSION REPRESSES ENDOGENOUS IKKBETA. PHOSPHORYLATION OF THE IKKBETA TARGET IKAPPABALPHA AND EXPRESSION OF KNOWN NF-KAPPAB TARGET GENES WERE IMPAIRED BY MIR-708. FURTHERMORE, WE IDENTIFIED AN ENHANCER REGION DOWNSTREAM OF THE MIR-708 PROMOTER THAT DISPLAYS A DISTINCT DNA METHYLATION STATUS IN CLL. HIGH ENHANCER METHYLATION IS SIGNIFICANTLY CORRELATED WITH LOWER MIR-708 EXPRESSION AND IS PREDOMINANTLY FOUND IN PATIENTS WITH POOR PROGNOSIS AND SHORTER TIME TO TREATMENT. THESE RESULTS DEMONSTRATE THAT MIR-708 REGULATES THE NF-KAPPAB PATHWAY BY TARGETING IKKBETA, AND THAT METHYLATION OF A KEY ENHANCER REGION CONTRIBUTES TO ITS SUPPRESSION IN CLL.	2015	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           
11 1966  37 EPIGENETIC ALTERATION OF PRKCDBP IN COLORECTAL CANCERS AND ITS IMPLICATION IN TUMOR CELL RESISTANCE TO TNFALPHA-INDUCED APOPTOSIS. PURPOSE: PRKCDBP IS A PUTATIVE TUMOR SUPPRESSOR IN WHICH ALTERATION HAS BEEN OBSERVED IN SEVERAL HUMAN CANCERS. WE INVESTIGATED EXPRESSION AND FUNCTION OF PRKCDBP IN COLORECTAL CELLS AND TISSUES TO EXPLORE ITS CANDIDACY AS A SUPPRESSOR IN COLORECTAL TUMORIGENESIS. EXPERIMENTAL DESIGN: EXPRESSION AND METHYLATION STATUS OF PRKCDBP AND ITS EFFECT ON TUMOR GROWTH WERE EVALUATED. TRANSCRIPTIONAL REGULATION BY NF-KAPPAB SIGNALING WAS DEFINED BY LUCIFERASE REPORTER AND CHROMATIN IMMUNOPRECIPITATION ASSAYS. RESULTS: PRKCDBP EXPRESSION WAS HARDLY DETECTABLE IN 29 OF 80 (36%) PRIMARY TUMORS AND 11 OF 19 (58%) CELL LINES, AND ITS ALTERATION CORRELATED WITH TUMOR STAGE AND GRADE. PROMOTER HYPERMETHYLATION WAS COMMONLY FOUND IN CANCERS. PRKCDBP EXPRESSION INDUCED THE G(1) CELL-CYCLE ARREST AND INCREASED CELLULAR SENSITIVITY TO VARIOUS APOPTOTIC STRESSES. PRKCDBP WAS INDUCED BY TNFALPHA, AND ITS LEVEL CORRELATED WITH TUMOR CELL SENSITIVITY TO TNFALPHA-INDUCED APOPTOSIS. PRKCDBP INDUCTION BY TNFALPHA WAS DISRUPTED BY BLOCKING NF-KAPPAB SIGNALING WHILE IT WAS ENHANCED BY RELA TRANSFECTION. THE PRKCDBP PROMOTER ACTIVITY WAS INCREASED IN RESPONSE TO TNFALPHA, AND THIS RESPONSE WAS ABOLISHED BY DISRUPTION OF A KAPPAB SITE IN THE PROMOTER. PRKCDBP DELAYED THE FORMATION AND GROWTH OF XENOGRAFT TUMORS AND IMPROVED TUMOR RESPONSE TO TNFALPHA-INDUCED APOPTOSIS. CONCLUSIONS: PRKCDBP IS A PROAPOPTOTIC TUMOR SUPPRESSOR WHICH IS COMMONLY ALTERED IN COLORECTAL CANCER BY PROMOTER HYPERMETHYLATION, AND ITS GENE TRANSCRIPTION IS DIRECTLY ACTIVATED BY NF-KAPPAB IN RESPONSE TO TNFALPHA. THIS SUGGESTS THAT PRKCDBP INACTIVATION MAY CONTRIBUTE TO TUMOR PROGRESSION BY REDUCING CELLULAR SENSITIVITY TO TNFALPHA AND OTHER STRESSES, PARTICULARLY UNDER CHRONIC INFLAMMATORY MICROENVIRONMENT.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        
12 1334  35 DEREGULATION OF AIOLOS EXPRESSION IN CHRONIC LYMPHOCYTIC LEUKEMIA IS ASSOCIATED WITH EPIGENETIC MODIFICATIONS. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS CHARACTERIZED BY A CLONAL ACCUMULATION OF MATURE NEOPLASTIC B CELLS THAT ARE RESISTANT TO APOPTOSIS. AIOLOS, A MEMBER OF THE IKAROS FAMILY OF ZINC-FINGER TRANSCRIPTION FACTORS, PLAYS AN IMPORTANT ROLE IN THE CONTROL OF MATURE B LYMPHOCYTE DIFFERENTIATION AND MATURATION. IN THIS STUDY, WE SHOWED THAT AIOLOS EXPRESSION IS UP-REGULATED IN B-CLL CELLS. THIS OVEREXPRESSION DOES NOT IMPLICATE ISOFORM IMBALANCE OR DISTURB AIOLOS SUBCELLULAR LOCALIZATION. THE CHROMATIN STATUS AT THE AIOLOS PROMOTER IN CLL IS DEFINED BY THE DEMETHYLATION OF DNA AND AN ENRICHMENT OF EUCHROMATIN ASSOCIATED HISTONE MARKERS, SUCH AS THE DIMETHYLATION OF THE LYSINE 4 ON HISTONE H3. THESE EPIGENETIC MODIFICATIONS SHOULD ALLOW ITS UPSTREAM EFFECTORS, SUCH AS NUCLEAR FACTOR-KAPPAB, CONSTITUTIVELY ACTIVATED IN CLL, TO GAIN ACCESS TO PROMOTER, RESULTING UP-REGULATION OF AIOLOS. TO DETERMINE THE CONSEQUENCES OF AIOLOS DEREGULATION IN CLL, WE ANALYZED THE EFFECTS OF AIOLOS OVEREXPRESSION OR DOWN-REGULATION ON APOPTOSIS. AIOLOS IS INVOLVED IN CELL SURVIVAL BY REGULATING THE EXPRESSION OF SOME BCL-2 FAMILY MEMBERS. OUR RESULTS STRONGLY SUGGEST THAT AIOLOS DEREGULATION BY EPIGENETIC MODIFICATIONS MAY BE A HALLMARK OF CLL.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        
13 2316  41 EPIGENETIC REGULATION OF FRUCTOSE-1,6-BISPHOSPHATASE 1 BY HOST TRANSCRIPTION FACTOR SPECKLED 110 KDA DURING HEPATITIS B VIRUS INFECTION. HEPATITIS B VIRUS (HBV) IS THE LEADING CAUSE OF LIVER DISEASE RANGING FROM ACUTE AND CHRONIC HEPATITIS TO LIVER CIRRHOSIS AND HEPATOCELLULAR CARCINOMA (HCC). STUDIES HAVE REVEALED THAT HBV INFECTION BROADLY REPROGRAMMES THE HOST CELLULAR METABOLIC PROCESSES FOR VIRAL PATHOGENESIS. PREVIOUS REPORTS HAVE SHOWN THAT GLYCOLYSIS AND GLUCONEOGENESIS ARE AMONG THE MOST DEREGULATED PATHWAYS DURING HBV INFECTION. WE NOTED THAT DESPITE BEING ONE OF THE RATE-LIMITING ENZYMES OF GLUCONEOGENESIS, THE ROLE AND REGULATION OF FRUCTOSE-1,6-BISPHOSPHATASE 1 (FBP1) DURING HBV INFECTION IS NOT MUCH EXPLORED. IN THIS STUDY, WE REPORT FBP1 UPREGULATION UPON HBV INFECTION AND UNRAVEL A NOVEL MECHANISM OF EPIGENETIC REPROGRAMMING OF FBP1 BY HBV VIA UTILIZING HOST FACTOR SPECKLED 110 KDA (SP110). HERE, WE IDENTIFIED ACETYLATED LYSINE 18 OF HISTONE H3 (H3K18AC) AS A SELECTIVE INTERACTOR OF SP110 BROMODOMAIN. FURTHERMORE, WE FOUND THAT SP110 GETS RECRUITED ON H3K18AC-ENRICHED FBP1 PROMOTER, AND FACILITATES RECRUITMENT OF DEACETYLASE SIRTUIN 2 (SIRT2) ON THAT SITE IN THE PRESENCE OF HBV. SIRT2 IN TURN BRINGS ITS INTERACTOR AND TRANSCRIPTIONAL ACTIVATOR HEPATOCYTE NUCLEAR FACTOR 4-ALPHA TO THE PROMOTER, WHICH ULTIMATELY LEADS TO A LOSS OF DNA METHYLATION NEAR THE COGNATE SITE. INTERESTINGLY, THIS SP110 DRIVEN FBP1 REGULATION DURING INFECTION WAS FOUND TO PROMOTE VIRAL-BORNE HCC PROGRESSION. MOREOVER, SP110 CAN BE USED AS A PROGNOSTIC MARKER FOR THE HEPATITIS-MEDIATED HCC PATIENTS, WHERE HIGH SP110 EXPRESSION SIGNIFICANTLY LOWERED THEIR SURVIVAL. THUS, THE EPIGENETIC READER PROTEIN SP110 HAS POTENTIAL TO BE A THERAPEUTIC TARGET TO CHALLENGE HBV-INDUCED HCCS.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            
14 6385  35 THE ROLE OF PROMYELOCYTIC LEUKEMIA PROTEIN IN STEATOSIS-ASSOCIATED HEPATIC TUMORS RELATED TO CHRONIC HEPATITIS B VIRUS INFECTION. THE PERSISTENCE OF HEPATITIS B SURFACE ANTIGEN (HBSAG) IS A RISK FACTOR FOR THE DEVELOPMENT OF STEATOSIS-ASSOCIATED TUMORS IN CHRONIC HEPATITIS B VIRUS (HBV) INFECTION, YET LITTLE IS KNOWN ABOUT THE METABOLIC LINK WITH THIS FACTOR. WE CORRELATED HBV-RELATED PATHOGENESIS IN GENETICALLY ENGINEERED MICE AND HUMAN CARRIERS WITH METABOLIC PROTEOMICS AND LIPOGENIC GENE EXPRESSION PROFILES. THE IMMUNOHISTOCHEMISTRY SHOWED THAT THE PROMYELOCYTIC LEUKEMIA PROTEIN (PML, A TUMOR SUPPRESSOR INVOLVED IN GENOME MAINTENANCE AND FATTY ACID OXIDATION), BEING INVERSELY INFLUENCED BY THE DYNAMIC HBSAG LEVELS FROM ACUTE PHASE TO SEROCLEARANCE, APPEARED AS A LIPO-METABOLIC SWITCH LINKING HBSAG-INDUCED STEATOSIS (LIPOGENESIS) TO HBSAG-LOST FAT-BURNING HEPATOCARCINOGENESIS (LIPOLYSIS). KNOCKDOWN OF PML IN HBSAG-TRANSGENIC MICE PREDISPOSED TO OBESITY AND DROVE EARLY STEATOSIS-SPECIFIC LIVER TUMORIGENESIS. PROTEOME ANALYSIS REVEALED THAT THE SIGNALING PATHWAYS CORRESPONDING TO ENERGY METABOLISM AND ITS REGULATORS WERE FREQUENTLY ALTERED BY SUPPRESSION OR DEPLETION OF PML IN THE HBSAG-TRANSGENIC MICE, MAINLY INCLUDING OXIDATIVE PHOSPHORYLATION AND FATTY ACID METABOLISM. EXPRESSION PROFILING FURTHER IDENTIFIED UPREGULATION OF STEAROYL-COA DESATURASE 1 (SCD1) AND EPIGENETIC METHYLATION OF NDUFA13 IN THE MITOCHONDRIAL RESPIRATORY CHAIN AND THE CELL CYCLE INHIBITOR CDKN1C IN CONCORDANCE TO THE INCREASED SEVERITY OF LIPODYSTROPHY AND NEOPLASIA IN THE LIVERS OF HBSAG-TRANSGENIC MICE WITH PML INSUFFICIENCY. THE DEFECT IN LIPOLYSIS IN PML-DEFICIENT HBSAG-TRANSGENIC MICE MADE THE HBSAG-INDUCED ADIPOSE-LIKE LIVER TUMORS VULNERABLE TO SYNTHETIC LETHALITY FROM TOXIC SATURATED FAT ACCUMULATION WITH A SCD1 INHIBITOR. OUR FINDINGS PROVIDE MECHANISTIC INSIGHTS INTO THE EVOLUTION OF STEATOSIS-ASSOCIATED HEPATIC TUMORS DRIVEN BY RECIPROCAL INTERACTIONS OF HBSAG AND PML, AND A POTENTIAL UTILITY OF LIPID METABOLIC REPROGRAMMING AS A TREATMENT TARGET.	2018	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         
15 6294  34 THE PROINFLAMMATORY CYTOKINE TNFALPHA INDUCES DNA DEMETHYLATION-DEPENDENT AND -INDEPENDENT ACTIVATION OF INTERLEUKIN-32 EXPRESSION. IL-32 IS A CYTOKINE INVOLVED IN PROINFLAMMATORY IMMUNE RESPONSES TO BACTERIAL AND VIRAL INFECTIONS. HOWEVER, THE ROLE OF EPIGENETIC EVENTS IN THE REGULATION OF IL-32 GENE EXPRESSION IS UNDERSTUDIED. HERE WE SHOW THAT IL-32 IS REPRESSED BY DNA METHYLATION IN HEK293 CELLS. USING CHIP SEQUENCING, LOCUS-SPECIFIC METHYLATION ANALYSIS, CRISPR/CAS9-MEDIATED GENOME EDITING, AND RT-QPCR (QUANTITATIVE RT-PCR) AND IMMUNOBLOT ASSAYS, WE FOUND THAT SHORT-TERM TREATMENT (A FEW HOURS) WITH THE PROINFLAMMATORY CYTOKINE TUMOR NECROSIS FACTOR ALPHA (TNFALPHA) ACTIVATES IL-32 IN A DNA DEMETHYLATION-INDEPENDENT MANNER. IN CONTRAST, PROLONGED TNFALPHA TREATMENT (SEVERAL DAYS) INDUCED DNA DEMETHYLATION AT THE PROMOTER AND A CPG ISLAND IN THE IL-32 GENE IN A TET (TEN-ELEVEN TRANSLOCATION) FAMILY ENZYME- AND NF-KAPPAB-DEPENDENT MANNER. NOTABLY, THE HYPOMETHYLATION STATUS OF TRANSCRIPTIONAL REGULATORY ELEMENTS IN IL-32 WAS MAINTAINED FOR A LONG TIME (SEVERAL WEEKS), CAUSING ELEVATED IL-32 EXPRESSION EVEN IN THE ABSENCE OF TNFALPHA. CONSIDERING THAT IL-32 CAN, IN TURN, INDUCE TNFALPHA EXPRESSION, WE SPECULATE THAT SUCH FEEDFORWARD EVENTS MAY CONTRIBUTE TO THE TRANSITION FROM AN ACUTE INFLAMMATORY RESPONSE TO CHRONIC INFLAMMATION.	2019	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
16 3049  33 GENOME-WIDE ANALYSIS REVEALS ZINC TRANSPORTER ZIP9 REGULATED BY DNA METHYLATION PROMOTES RADIATION-INDUCED SKIN FIBROSIS VIA THE TGF-BETA SIGNALING PATHWAY. RADIATION-INDUCED SKIN FIBROSIS IS A DETRIMENTAL AND CHRONIC DISORDER THAT OCCURS AFTER RADIATION EXPOSURE. DNA METHYLATION HAS BEEN CHARACTERIZED AS AN IMPORTANT REGULATORY MECHANISM OF MULTIPLE PATHOLOGICAL PROCESSES. IN THIS STUDY, WE COMPARED THE GENOME-WIDE DNA METHYLATION STATUS IN RADIATION-INDUCED FIBROTIC SKIN AND ADJACENT NORMAL TISSUES OF RATS BY METHYLATED DNA IMMUNOPRECIPITATION SEQUENCING. RADIATION-INDUCED FIBROTIC SKIN SHOWED DIFFERENTIALLY METHYLATED REGIONS ASSOCIATED WITH 3,650 PROTEIN-CODING GENES, 72 MICRORNAS, 5,836 LONG NONCODING RNAS AND 3 PIWI-INTERACTING RNAS. BY INTEGRATING THE MRNA AND METHYLATION PROFILES, THE ZINC TRANSPORTER SLC39A9/ZIP9 WAS INVESTIGATED IN GREATER DETAIL. THE PROTEIN LEVEL OF ZIP9 WAS INCREASED IN IRRADIATED SKIN TISSUES OF HUMANS, MONKEYS, AND RATS, ESPECIALLY IN RADIOGENIC FIBROTIC SKIN TISSUES. RADIATION INDUCED THE DEMETHYLATION OF A CPG DINUCLEOTIDE IN EXON 1 OF ZIP9 THAT RESULTED IN RECRUITMENT OF THE TRANSCRIPTIONAL FACTOR SP1 AND INCREASED ZIP9 EXPRESSION. OVEREXPRESSION OF ZIP9 RESULTED IN ACTIVATION OF THE PROFIBROTIC TRANSFORMING GROWTH FACTOR-BETA SIGNALING PATHWAY THROUGH PROTEIN KINASE B IN HUMAN FIBROBLASTS. IN ADDITION, RADIATION-INDUCED SKIN FIBROSIS WAS ASSOCIATED WITH INCREASED ZINC ACCUMULATION. THE ZINC CHELATOR N,N,N',N'-TETRAKIS(2-PYRIDYLMETHYL)-1,2-ETHYLENEDIAMINE ABROGATED ZIP9-INDUCED ACTIVATION OF THE TRANSFORMING GROWTH FACTOR-BETA SIGNALING PATHWAY AND ATTENUATED RADIATION-INDUCED SKIN FIBROSIS IN A RAT MODEL. IN SUMMARY, OUR FINDINGS ILLUSTRATE EPIGENETIC REGULATION OF ZIP9 AND ITS CRITICAL ROLE IN PROMOTING RADIATION-INDUCED SKIN FIBROSIS.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   
17 4233  33 METHYLATION OF SEPTIN9 MEDIATED BY DNMT3A ENHANCES HEPATIC STELLATE CELLS ACTIVATION AND LIVER FIBROGENESIS. LIVER FIBROSIS, RESULTING FROM CHRONIC AND PERSISTENT INJURY TO THE LIVER, IS A WORLDWIDE HEALTH PROBLEM. ADVANCED LIVER FIBROSIS RESULTS IN CIRRHOSIS, LIVER FAILURE AND EVEN HEPATOCELLULAR CANCER (HCC), OFTEN EVENTUALLY REQUIRING LIVER TRANSPLANTATION, POSES A HUGE HEALTH BURDEN ON THE GLOBAL COMMUNITY. HOWEVER, THE SPECIFIC PATHOGENESIS OF LIVER FIBROSIS REMAINS NOT FULLY UNDERSTOOD. NUMEROUS BASIC AND CLINICAL STUDIES HAVE PROVIDED EVIDENCE THAT EPIGENETIC MODIFICATIONS, ESPECIALLY DNA METHYLATION, MIGHT CONTRIBUTE TO THE ACTIVATION OF HEPATIC STELLATE CELLS (HSCS), THE PIVOTAL CELL TYPE RESPONSIBLE FOR THE FIBROUS SCAR IN LIVER. HERE, REDUCED REPRESENTATION BISULFITE SEQUENCING (RRBS) AND BISULFITE PYROSEQUENCING PCR (BSP) ANALYSIS IDENTIFIED HYPERMETHYLATION STATUS OF SEPTIN9 (SEPT9) GENE IN LIVER FIBROGENESIS. SEPT9 PROTEIN WAS DRAMATICALLY DECREASED IN LIVERS OF CCL4-TREATED MICE AND IMMORTALIZED HSC-T6 CELLS EXPOSED TO TGF-BETA1. NEVERTHELESS, THE SUPPRESSION OF SEPT9 COULD BE BLOCKED BY DNMT3A-SIRNA AND DNA METHYLTRANSFERASE INHIBITOR, 5-AZA-2'-DEOXYCYTIDINE (5-AZADC). OVEREXPRESSED SEPT9 ATTENUATED TGF-BETA1-INDUCED EXPRESSION OF MYOFIBROBLAST MARKERS ALPHA-SMA AND COL1A1, ACCOMPANIED BY UP-REGULATION OF CELL APOPTOSIS-RELATED PROTEINS. CONVERSELY, RNAI-MEDIATED SILENCING OF SEPT9 ENHANCED ACCUMULATION OF EXTRACELLULAR MATRIX. THESE OBSERVATIONS SUGGESTED THAT SEPT9 CONTRIBUTED TO ALLEVIATE LIVER FIBROSIS MIGHT PARTIALLY THROUGH PROMOTING ACTIVATED HSCS APOPTOSIS AND THIS ANTI-FIBROGENESIS EFFECT MIGHT BE BLOCKED BY DNMT-3A MEDIATED METHYLATION OF SEPT9. THEREFORE, PHARMACOLOGICAL AGENTS THAT INHIBIT SEPT9 METHYLATION AND INCREASE ITS EXPRESSION COULD BE CONSIDERED AS VALUABLE TREATMENTS FOR LIVER FIBROSIS.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              
18  155  28 ABERRANT METHYLATION OF POLO-LIKE KINASE CPG ISLANDS IN PLK4 HETEROZYGOUS MICE. BACKGROUND: HEPATOCELLULAR CARCINOMA (HCC), ONE OF THE MOST COMMON CANCERS WORLD-WIDE OCCURS TWICE AS OFTEN IN MEN COMPARED TO WOMEN. PREDISPOSING CONDITIONS SUCH AS ALCOHOLISM, CHRONIC VIRAL HEPATITIS, AFLATOXIN B1 INGESTION, AND CIRRHOSIS ALL CONTRIBUTE TO THE DEVELOPMENT OF HCC. METHODS: WE USED A COMBINATION OF METHYLATION SPECIFIC PCR AND BISULFITE SEQUENCING, QREAL-TIME PCR (QPCR), AND WESTERN BLOT ANALYSIS TO EXAMINE EPIGENETIC CHANGES FOR THE POLO-LIKE KINASES (PLKS) DURING THE DEVELOPMENT OF HEPATOCELLULAR CARCINOMA (HCC) IN PLK4 HETEROZYGOUS MICE AND MURINE EMBRYONIC FIBROBLASTS (MEFS). RESULTS: HERE WE REPORT THAT THE PROMOTER METHYLATION OF PLK4 CPG ISLANDS INCREASES WITH AGE, WAS MORE PREVALENT IN MALES AND THAT PLK4 EPIGENETIC MODIFICATION AND SUBSEQUENT DOWNREGULATION OF EXPRESSION WAS ASSOCIATED WITH THE DEVELOPMENT OF HCC IN PLK4 MUTANT MICE. INTERESTINGLY, THE OPPOSITE OCCURS WITH ANOTHER PLK FAMILY MEMBER, PLK1 WHICH WAS TYPICALLY HYPERMETHYLATED IN NORMAL LIVER TISSUE BUT BECAME HYPOMETHYLATED AND UPREGULATED IN LIVER TUMOURS. FURTHERMORE, UPON ALCOHOL EXPOSURE MURINE EMBRYONIC FIBROBLASTS EXHIBITED INCREASED PLK4 HYPERMETHYLATION AND DOWNREGULATION ALONG WITH INCREASED CENTROSOME NUMBERS AND MULTINUCLEATION. CONCLUSIONS: THESE RESULTS SUGGEST THAT ABERRANT PLK METHYLATION IS CORRELATED WITH THE DEVELOPMENT OF HCC IN MICE.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         
19 5004  31 PERIPHERAL B CELLS FROM PATIENTS WITH HEPATITIS C VIRUS-ASSOCIATED LYMPHOMA EXHIBIT CLONAL EXPANSION AND AN ANERGIC-LIKE TRANSCRIPTIONAL PROFILE. CHRONIC HCV INFECTION REMAINS A GLOBAL HEALTH CONCERN DUE TO ITS INVOLVEMENT IN HEPATIC AND EXTRAHEPATIC DISEASES, INCLUDING B CELL NON-HODGKIN LYMPHOMA (BNHL). CLINICAL AND EPIDEMIOLOGICAL EVIDENCE SUPPORT A CAUSAL ROLE FOR HCV IN BNHL DEVELOPMENT, ALTHOUGH MECHANISTIC INSIGHT IS LACKING. WE PERFORMED RNA-SEQUENCING ON PERIPHERAL B CELLS FROM PATIENTS WITH HCV ALONE, BNHL ALONE, AND HCV-ASSOCIATED BNHL TO IDENTIFY UNIQUE AND SHARED TRANSCRIPTIONAL PROFILES ASSOCIATED WITH TRANSFORMATION. IN PATIENTS WITH HCV-ASSOCIATED BNHL, WE OBSERVED THE ENRICHMENT OF AN ANERGIC-LIKE GENE SIGNATURE AND EVIDENCE OF CLONAL EXPANSION THAT WAS CORRELATED WITH THE EXPRESSION OF EPIGENETIC REGULATORY GENES. OUR DATA SUPPORT A ROLE FOR VIRAL-MEDIATED CLONAL EXPANSION OF ANERGIC-LIKE B CELLS IN HCV-ASSOCIATED BNHL DEVELOPMENT AND SUGGEST EPIGENETIC DYSREGULATION AS A POTENTIAL MECHANISM DRIVING EXPANSION. WE PROPOSE EPIGENETIC MECHANISMS MAY BE INVOLVED IN BOTH HCV-ASSOCIATED LYMPHOMA AND REGULATION OF B CELL ANERGY, REPRESENTING AN ATTRACTIVE TARGET FOR CLINICAL INTERVENTIONS.	2023	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                          
20 3247  41 HEPATITIS B VIRUS BASAL CORE PROMOTER MUTATIONS SHOW LOWER REPLICATION FITNESS ASSOCIATED WITH CCCDNA ACETYLATION STATUS. IN CHRONIC HEPATITIS B VIRUS (HBV) INFECTION, VARIANTS WITH MUTATIONS IN THE BASAL CORE PROMOTER (BCP) AND PRECORE REGION PREDOMINATE AND ASSOCIATE WITH MORE SEVERE DISEASE FORMS. STUDIES ON THEIR EFFECT ON VIRAL REPLICATION REMAIN CONTROVERSIAL. INCREASING EVIDENCE SHOWS THAT EPIGENETIC MODIFICATIONS OF CCCDNA REGULATE HBV REPLICATION AND DISEASE OUTCOME. HERE WE DETERMINED THE TRANSCRIPTION AND VIRAL REPLICATION EFFICIENCY OF WELL-DEFINED BCP AND PRECORE MUTATIONS AND THEIR EFFECT ON CCCDNA EPIGENETIC CONTROL. HBV MONOMERS BEARING BCP MUTATIONS A1762T/G1764A AND A1762T/G1764A/C1766T, AND PRECORE MUTATIONS G1896A, G1899A AND G1896A/G1899A, WERE TRANSFECTED INTO HEPG2 CELLS USING A PLASMID-FREE APPROACH. VIRAL RNA TRANSCRIPTS WERE DETECTED BY NORTHERN BLOT HYBRIDIZATION AND RT PCR, DNA REPLICATIVE INTERMEDIATES BY SOUTHERN BLOTTING AND RT PCR, AND VIRAL RELEASE WAS MEASURED BY ELISA. ACETYLATION OF CCCDNA-BOUND HISTONES WAS ASSESSED BY CHROMATIN IMMUNOPRECIPITATION (CHIP) ASSAY AND METHYLATION OF CCCDNA BY BISULFITE SEQUENCING. BCP MUTATIONS RESULTED IN LOW VIRAL RELEASE, MRNA TRANSCRIPTION AND PGRNA/CCCDNA RATIOS THAT PARALLELED THE ACETYLATION OF CCCDNA-BOUND H4 HISTONE AND INVERSELY CORRELATED WITH THE HDAC1 RECRUITMENT ONTO CCCDNA. INDEPENDENTLY OF THE MUTATIONS, CCCDNA WAS A TARGET FOR METHYLATION, ACCOMPANIED BY THE UPREGULATION OF DNMT1 EXPRESSION AND DNMT1 RECRUITMENT ONTO CCCDNA. OUR RESULTS SUGGEST THAT BCP MUTATIONS DECREASE VIRAL REPLICATION CAPACITY POSSIBLY BY MODULATING THE ACETYLATION AND DEACETYLATION OF CCCDNA-BOUND HISTONES WHILE PRECORE MUTATIONS DO NOT HAVE A SIGNIFICANT EFFECT ON VIRAL REPLICATION. THESE DATA PROVIDE EVIDENCE THAT EPIGENETIC FACTORS CONTRIBUTE TO THE REGULATION OF HBV VIRAL REPLICATION.	2016