title: Nuclear redox imbalance affects circadian oscillation in HaCaT keratinocytes.
abstract: Circadian clock is regulated by a transcriptional/translational feedback loop (TTFL) lasting ∼24 h. Circadian oscillation of peroxiredoxins (PRDX1-6) redox status has been shown in mature erythrocytes. We have recently reported that nuclear levels of PRDX2 are circadian regulated in the HaCaT keratinocytes. In this study, we addressed whether PRDX2 translocation could influence the TTFL. A reporter HaCaT cell line stably expressing the luciferase gene under control of Bmal1 promoter was lentivirally transduced either with an empty vector (EV), a vector carrying a myc-tagged wild type PRDX2 (PRDX2-Myc) or the same gene with a nuclear localization sequence (PRDX2-MycNuc). PRDX2 overexpressing cells were protected from H2O2-induced oxidative stress. The amplitude of the Bmal1 promoter activity was significantly dampened in PRDX2-MycNuc versus EV cells when synchronized either by dexamethasone treatment or temperature cycles. Clock synchronization was not affected in PRDX2 silenced cells. N-acetyl cysteine or melatonin treatments, significantly dampened the Bmal1 promoter activity suggesting that sustained scavenging of ROS impairs clock synchronization. Noteworthy, H2O2 treatment rescued proper oscillation of the clock in synchronized PRDX2-MycNuc HaCaT cells. Since the histone deacetylase Sirtuin 1 (Sirt1) modulates clock gene expression amplitude, the effect of Sirt1 activator resveratrol or Sirt1 inhibitor nicotinamide were also investigated. Interestingly, NAM enhanced the molecular clock synchronization in PRDX2-MycNuc cells. Our findings demonstrate that PRDX2 regulates the TTFL oscillation by finely tuning the cellular redox status of the nucleus likely influencing the deacetilase activity of SIRT1 enzyme.
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Medicina Clinica e Molecolare, Sapienza Università di Roma, Rome, Italy.
authors.forename: Danilo
authors.initials: D
authors.lastname: Ranieri
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Medicina Clinica e Molecolare, Sapienza Università di Roma, Rome, Italy; Laboratorio di Biologia Vascolare e Medicina Rigenerativa, Centro Cardiologico Monzino IRCCS, Milan, Italy. Electronic address: daniele.avitabile@uniroma1.it.
authors.forename: Daniele
authors.initials: D
authors.lastname: Avitabile
authors.affiliation: Department of Urology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Shiota
authors.affiliation: Department of Urology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Yokomizo
authors.affiliation: Department of Urology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Seiji
authors.initials: S
authors.lastname: Naito
authors.affiliation: Dipartimento di Medicina Sperimentale, Sapienza Università di Roma, Italy.
authors.forename: Mariano
authors.initials: M
authors.lastname: Bizzarri
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Medicina Clinica e Molecolare, Sapienza Università di Roma, Rome, Italy; Azienda Ospedaliera S. Andrea, Rome, Italy.
authors.forename: Maria Rosaria
authors.initials: MR
authors.lastname: Torrisi
mesh_terms: D056930:ARNTL Transcription Factors / Q000235:genetics / Q000378:metabolism; D000111:Acetylcysteine / Q000494:pharmacology; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002940:Circadian Rhythm / Q000187:drug effects / Q000502:physiology*; D015870:Gene Expression; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008550:Melatonin / Q000494:pharmacology; D009536:Niacinamide / Q000494:pharmacology; D010084:Oxidation-Reduction; D054464:Peroxiredoxins / Q000096:biosynthesis / Q000378:metabolism; D014176:Protein Biosynthesis; D017382:Reactive Oxygen Species; D000077185:Resveratrol; D056564:Sirtuin 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology; D014158:Transcription, Genetic
chemical_list: D056930:ARNTL Transcription Factors; C538960:BMAL1 protein, human; D017382:Reactive Oxygen Species; D013267:Stilbenes; D009536:Niacinamide; C086065:PRDX2 protein, human; D054464:Peroxiredoxins; C447939:SIRT1 protein, human; D056564:Sirtuin 1; D008550:Melatonin; D000077185:Resveratrol; D000111:Acetylcysteine
keywords: Circadian rhythms; HaCaT keratinocytes; Melatonin; N-acetyl cysteine; Nicotinamide; Oxidative stress; Peroxiredoxin 2; Resveratrol; Sirt1