Circadian Genes Per2 Research Articles

Rotating-shift nurses after a day off: peripheral clock gene expression, urinary melatonin, and serum 17-β-estradiol levels

Impairment of clock gene expression and changes in melatonin and 17-β-estradiol levels may constitute biological alterations underlying the increased risk of breast cancer among shift workers. The aim of this study was to compare levels of selected core clock gene expression, 6-sulfatoxymelatonin (aMT6s), and 17-β-estradiol between rotational shift work (SW) and daytime (DT) workers after a day off. The cross-sectional study comprised 60 nurses with ≥2 years of SW and 56 permanent DT nurses. Transcript levels of circadian genes BMAL1, CLOCK, NPAS2, CRY1, CRY2, PER1, PER2, PER3, and REVERBα were determined by quantitative real-time polymerase chain reaction (PCR) in lymphocytes. All participants were tested in the early follicular phase of the menstrual cycle. Samples were collected at the beginning of the morning-shift after a regular night's sleep on a day off. Chronotype and sociodemographic characteristics were also evaluated. We found a significantly higher expression of BMAL1, CLOCK, NPAS2, PER1, PER2, and REVERBα and a lower expression of PER3, CRY1 and CRY2 among SW compared to DT nurses. SW participants did not demonstrate a significant difference in aMT6s levels, but they did show significantly higher 17-β-estradiol levels compared to DT nurses. Multiple linear regression analysis confirmed the role of SW on expression of BMAL1 (β 0.21, P=0.040), CLOCK (β 0.35, P=0.008), NPAS2 (β 0.30, P=0.012), PER1 (β 0.33, P=0.008), PER2 (β 0.19, P=0.047), PER3 (β -0.27, P=0.012), CRY1 (β -0.33, P=0.002), CRY2 (β -0.31, P=0.005), REVERBα (β 0.19, P=0.045), and on 17-β-estradiol levels (β 0.32, P=0.003). The analysis also confirmed the role of chronotype as an independent factor for PER1 (β 0.48, P=0.001) and PER2 (β -0.22, P=0.022) expression, and 17-β-estradiol levels (β 0.26, P=0.011). Rotating SW nurses show alterations in peripheral clock gene expression and 17-β-estradiol levels at the beginning of the morning shift after a day off.

Deletion of circadian gene Per1 alleviates acute ethanol-induced hepatotoxicity in mice

The severity of ethanol-induced liver injury is associated with oxidative stress and lipid accumulation in the liver. Core circadian clock is known to mediate antioxidative enzyme activity and lipid metabolism. However, the link between circadian clock and ethanol-induced hepatotoxicity remains unclear. Here we showed that extents of acute ethanol-induced liver injury and steatosis in mice exhibit circadian variations consistent with hepatic expression of Period (Per) genes. Mice lacking clock gene Per1 displayed less susceptible to ethanol-induced liver injury, as evidenced by lower serum transaminase activity and less severe histopathological changes. Ethanol-induced lipid peroxidation was alleviated in Per1−/− mice. However, Per1 deletion had no effect on antioxidants depletion caused by ethanol administration. Ethanol-induced triglycerides (TG) accumulation in the serum and liver was significantly decreased in Per1−/− mice compared with that in wild-type (WT) mice. Analysis of gene expression in the liver revealed peroxisome proliferators activated receptor-gamma (PPARγ) and its target genes related to TG synthesis are remarkably down-regulated in Per1−/− mice. HepG2 cells were treated with ethanol at 150mM for 3 days. Per1 overexpression augmented lipid accumulation after treatment with ethanol in HepG2 cells, but had no effect on ethanol-induced oxidative stress. Expression of genes related to lipogenesis, including PPARγ and its target genes, was up-regulated in cells overexpressing Per1. In conclusion, these results indicated that circadian rhythms of ethanol-induced hepatotoxicity are controlled by clock gene Per1, and deletion of Per1 protected mice from ethanol-induced liver injury by decreasing hepatic lipid accumulation.

The influence of L-arginine on circadian rhythm and circadian period genes

To study the effect of L-arginine on circadian system, ICR mice and AR4-2J cells were treated with L-arginine. The locomotor activity of the mice intraperitoneally injected with a high dose of L-arginine was observed. We also detected effects of arginine on Per1 and Per2 gene expressions in AR4-2J cells. Double-plotted actograms were applied to analyze the locomotor activity of the mice, and reverse transcription-polymerase chain reaction (RT-PCR) was used in analysis of the expression of the major circadian genes Per1 and Per2 in AR4-2J cells. The results presented here indicate that the intraperitoneal injection of L-arginine would strongly disrupt the circadian rhythm of locomotor activity in mice. While the co-culture of L-arginine would also disturb the circadian rhythm of Per1 and Per2 genes which were induced by PMA in AR4-2J cells, the expression levels of both genes greatly decreased.

Clinical research on radiotherapy at different time for unresectable cervical cancer.

e16516 Background: Radiotherapy is one of the mainstay treatments for cervical cancer. Circadian rhythm can be associated with cancers. In this study, we focused on the therapeutic effects, acute radiation damages and expressions of circadian genes in unresectable cervical cancer irradiated at different time. Methods: A total of 67 patients of unresectable cervical cancer (stage IIB-IIIB) were randomized to morning (9:00-11:00am) and evening (9:00-11:00pm) irradiation groups. Patients were treated with external beam radiotherapy to the whole pelvis (50 Gy in 25 fractions) and brachytherapy (36-42 Gy in 6-7 fractions). Therapeutic effects and acute radiation toxicities were evaluated after radiotherapy. Samples analyzed by flow cytometry were obtained from cervix at 9:00-11:00am and 9:00-11:00pm before and after radiotherapy. Results: Of 65 patients, the overall and higher grade of diarrhea (III-IV) induced by radiation in patients after morning vs evening radiotherapy were 75.0% vs 57.6% (p = 0.138), 12.5% vs 6.1% (P=0.427). Higher grade of hematologica toxicity in evening group was significantly increased as compared with morning group (54.5% vs 21.9%. p = 0.007). But patients’ response to radiation in two groups was similar (p = 0.708). Increased cell apoptosis and decreased G2/M phase fraction were observed significantly both in two groups after radiotherapy, but the increased/decreased values between two groups were found not to differ significantly. Expressions of circadian genes Per1, Per2 and Clock also increased after radiotherapy, and the increased levels between two groups were similar. Moreover, there were significant negative correlations between expressions of circadian genes Per1, Per2, Clock and G2/M phase fraction. But their expressions were positively correlated with cell apoptosis. Conclusions: Circadian genes may be associated with the similar radiation response and different toxicity in cervical cancer irradiated at different time by influencing cell cycle and apoptosis. The knowledge may provide a possibility of treating the patients with increased effect and decreased toxicity. Clinical trial information: ChiCTR-TRC-10001228.

Abstract 5303: Biomarker development for the evaluation of molecular circadian desynchrony in a single measurement.

Abstract Most organisms possess an endogenous circadian clock which is responsible for the temporal organization of rhythmic biochemical and metabolic processes. At the molecular level, rhythmicity in peripheral tissues is maintained by a core set of circadian genes, which in turn influence the expression programs of a large number of additional genes. Disruption in circadian rhythms has been implicated in a variety of human diseases, including cancer. However, the dynamic nature of the circadian system complicates study of molecular rhythms, and this is compounded by the inherent tissue-specificity of biological rhythms, which preclude the use of surrogate tissue. Since serial sampling of target peripheral tissue is unrealistic in human studies, our goal is to produce a model that will allow single-measurement evaluation of molecular desynchrony in archived tissue by identifying in-phase gene expression signatures in circadian gene pairs that vary over time on an absolute scale, but do not vary relative to one another. Using next-generation sequencing technologies, we have annotated the complete circadian transcriptome in primary, pre-stasis human mammary epithelial cells (HMECs) with sampling every 4-hours over a 24-hour time period. Cycling transcripts were identified using the non-parametric JTK_CYCLE algorithm, with further stratification according to circadian phase. In-phase transcript pairs were identified by calculating the standard error of the mean difference (SEM) in normalized gene expression across all time points for all pairwise comparisons among genes within each phase. Candidate pairs were then selected for further investigation, both in vitro and in normal and tumor tissue profiling experiments extracted from publicly available datasets, in order to evaluate their utility as biomarkers of tumor-associated desynchrony. Here we present the example of one such pair: IFNGR2 and PER1. In our primary HMEC model, IFNGR2 expression varies in a circadian manner over time, but maintains a consistent relationship with the core circadian gene PER1 with an approximately two-fold difference at each time-point (mean ratio = 2.29, SEM = 0.2280). In array data from single time point measurements of human mammary tissue (Array Express accession number E-TABM-276), the relationship between IFNGR2 and PER1 expression is similar to the tissue culture model in normal tissue, but becomes increasingly desynchronous in tissue with cystic changes (mean ratio = 3.83, SEM = 0.5717, P = 0.0422), and invasive carcinoma tissue (mean ratio=9.12, SEM = 0.9367, P = 0.0002). This data provides proof of principle that objective, quantitative biomarkers of molecular circadian disruption can be identified. In addition, these biomarkers can be useful for single measurement analysis, and are shown to correlate with the progression from normal tissue to invasive carcinoma. Citation Format: Tracy L. Mandichak, Joseph C. Combs, Elizabeth J. Ferree, Aaron E. Hoffman. Biomarker development for the evaluation of molecular circadian desynchrony in a single measurement. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5303. doi:10.1158/1538-7445.AM2013-5303

Expression and clinical significance of circadian gene Per2 in non-small cell lung cancer

To detect the expression of Per2 in non-small cell lung cancer (NSCLC), and analyze its clinical significance. The expression of Per2 was determined in 60 NSCLC and 20 normal lung tissues by immunohistochemical assay, and the relationship between Per2 expression and clinicopathological features was analyzed. The positive expression rates of Per2 in NSCLC and normal lung tissues were 71.7% and 95.0%, respectively (P < 0.05). The expression of Per2 in NSCLC was correlated with pathological differentiation and TNM stage (P < 0.05). The expression of Per2 in NSCLC is decreased. The negative expression of Per2 may contribute to the development and invasion in NSCLC.

The Circadian Gene Expression of Per1 and Per2 and Their Influence on Radiotherapeutic Sensitivity of Glioma In Vitro

Per2 plays a key role in regulating the circadian rhythm in mammals. However, the circadian clock gene expression of Per1 and Per2 and its influence on radiotherapeutic sensitivity of C6 glioma cells in vitro have not been explored. Aim: To investigate the rhythm expression of circadian gene Per1 and Per2, and examine the influence on radiotherapeutic sensitivity of two important clock genes in C6 glioma cells. Materials &amp; methods: The cultured C6 glioma cells and NIH3T3 cells were stimulated by phorbol 12-myristate 13-acetate (PMA). The expression of Per1 and Per2 at the indicated times were examined with a method for the absolute quantification of cDNA using real-time PCR. The cultured cell were given x-irradiation at the indicated times and the cell-cycle, apoptosis and proliferation were examined by flow cytometry. Results: We report here that PMA treatment of C6 rat glioma cells induces circadian expression of Per2, and that during periods of high expression, cells are blocked at the G2/M transition and are more sensitive to x-irradiation. PMA treatment of NIH3T3 cells induced circadian expression of Per1 and Per2, but high Per expression did not block the cell cycle or render the cells more sensitive to irradiation. Conclusion: Our results suggest that Per2 expression may increase the efficacy of radiotherapy against glioma.

Common transcriptional effects in the mouse striatum following chronic treatment with heroin and methamphetamine

The molecular alterations that underlie the long-lasting behavioural effects of drugs of abuse, such as psychomotor sensitization and physical dependence, are still not known. Moreover, it is not known which molecular effects are similar for addictive drugs from various pharmacological classes. In this study, we utilized whole-genome microarray profiling to evaluate the detailed time-course of transcriptional alterations in the mouse striatum during chronic treatment with heroin (HER) and methamphetamine (METH) and after period of spontaneous withdrawal. We identified 27 genes regulated by chronic drug administration. The overlap between lists of HER- and METH-induced genes was highly significant. The most substantial impact on the gene expression profile was observed for circadian genes (Per1, Per2 and Nr1d1). However, changing the treatment scheme from diurnal to nocturnal was sufficient to attenuate the drug-induced changes in circadian gene mRNA levels. Both of the drugs caused a dose-dependent induction in glucocorticoid-dependent genes with relatively long mRNA half-lives (Fkbp5, Sult1a1 and Plin4). The analysis also showed a drug-regulated group of transcripts enriched in the nucleus accumbens and includes well known (Pdyn, Cartpt and Rgs2) as well as new (Fam40b and Inmt) candidate genes. All identified alterations in the striatal transcriptome were transient and persisted up to 6 days after withdrawal. Behavioural sensitization, however, was maintained throughout the 12-day withdrawal period for both HER and METH. We suggest that transient gene expression alterations during drug treatment and in the early period of withdrawal are involved in the establishment of persistent neuroplastic alterations responsible for the development of drug addiction.

Expressions of the circadian genes Per2, Bmal1, Clock and Cry1 during the different stages of follicular development and their regulation by FSH in bovine granulosa cells from small follicles

Circadian clock genes are involved in mammalian reproductive physiology. We examined the expression of circadian clock genes such as Per2, Bmal1, Clock and Cry in granulosa cells from follicles at different developmental stages and in follicle-stimulating hormone (FSH)-treated granulosa cells. The expression of Clock mRNA was significantly increased in granulosa cells from pre-selection follicles as compared to that in the granulosa cells from other follicles. In contrast, the expression of Per2 mRNA was significantly increased in the granulosa cells from preovulatory follicles as compared to that in the granulosa cells from other follicles. Bmal1 and Cry mRNA expressions did not change across different follicular stages. FSH stimulated the expression of Clock mRNA in the cultured granulosa cells. The expression of Bmal1, Cry and Per2 mRNAs was not affected by FSH. Our results suggested that Clock may be associated with follicular development in the bovine ovary. In summary, our results suggested that circadian clock genes play an important role during follicular development in the bovine ovary.

Expression of circadian gene Per2 and VEGF in NSCL tissue and its clinical significance

Objective To detect the expression of Per2 and VEGF in NSCLC tissue,and analyze its clinical significance.Methods The expression of Per2 and VEGF was measured in 60 NSCLC and 20 normal lung tissue by immunohistochemistry assay,then the relation with clinicopathological parameters was analyzed.Results The positive expression rate of Per2 in NSCLC and in normal lung tissue were 71.7％ and 95.0％ ( x2 =4.683,P ＜0.05 ),while the positive expression rate of VEGF were 58.3％ and 15％ (x2 =11.295,P ＜ 0.01 ).The negative expression of Per2 in NSCLC was correlated with pathological differentiation or TNM stage( x2 =4.413,6.179,all P ＜0.05),while the positive expression of VEGF in NSCLC was correlated with pathological differentiation or lymphatic metastasis (x2 =6.524,P ＜ 0.05 ).The expression of Per2 was negatively correlated with the expression of VEGF in NSCLC (r =-0.381,P ＜ 0.01 ).Conclusion The overexpression of VEGF in NSCLC may contribute to invasion and metastasis,while Per2 may be a beneficial prognosis factor in patients with NSCLC. Key words: Cancer, non-small-cell lung; Circadian gene Per2; Vascular endothelial growth factor; Immunohistochemistry

Tissue-Specific Function of Period3 in Circadian Rhythmicity

The mammalian circadian system is composed of multiple central and peripheral clocks that are temporally coordinated to synchronize physiology and behavior with environmental cycles. Mammals have three homologs of the circadian Period gene (Per1, 2, 3). While numerous studies have demonstrated that Per1 and Per2 are necessary for molecular timekeeping and light responsiveness in the master circadian clock in the suprachiasmatic nuclei (SCN), the function of Per3 has been elusive. In the current study, we investigated the role of Per3 in circadian timekeeping in central and peripheral oscillators by analyzing PER2::LUCIFERASE expression in tissues explanted from C57BL/6J wild-type and Per3−/− mice. We observed shortening of the periods in some tissues from Per3−/− mice compared to wild-types. Importantly, the periods were not altered in other tissues, including the SCN, in Per3−/− mice. We also found that Per3-dependent shortening of endogenous periods resulted in advanced phases of those tissues, demonstrating that the in vitro phenotype is also present in vivo. Our data demonstrate that Per3 is important for endogenous timekeeping in specific tissues and those tissue-specific changes in endogenous periods result in internal misalignment of circadian clocks in Per3−/− mice. Taken together, our studies demonstrate that Per3 is a key player in the mammalian circadian system.

The Influence of Circadian Gene Per2 on Cell Damaged by Ultraviolet C

It has been shown that circadian genes not only play an important role on circadian rhythms, but also participate in other physiological and pathological activities, such as drug dependence, cancer development and radiation injury. The Per2, an indispensable component of the circadian clock, not only modulates circadian oscillations, but also regulates organic function. In the present study, we applied mPER2-upregulated NIH3T3 cells to reveal the relationship of mPer2 and the cells damaged by ultraviolet C (UVC). NIH3T3 cells at the peak of the expression of mPer2 induced by phorbol 12-myristate 13-acetate (PMA) demonstrated little damage by UVC evaluated by MTT assay, cell growth curves and cell colony-forming assay, compared with that at the nadir of the expression of mPer2. Overexpression of mPER2, accompanied p53 upregulated, also demonstrated protective effect on NIH3T3 cells damaged by UVC. These results suggest that mPer2 plays a protective effect on cells damaged by UVC, whose mechanism may be involved in upregulated p53.

Daily Rhythmic Expression Patterns of Clock1a, Bmal1, and Per1 Genes in Retina and Hypothalamus of the Rainbow Trout, Oncorhynchus Mykiss

Living organisms show daily rhythms in physiology, behavior, and gene expression, which are due to the presence of endogenous clocks that synchronize biological processes to the 24-h light/dark cycle. In metazoans, generation of circadian rhythmicity is a consequence of specialized tissues known as “master clocks,” having different locations among species. A few studies have described clock-gene expression in fish neural tissues, but none of them assessed clock-gene expression in different discrete regions. The present study was designed to explore the presence/absence of circadian clock-gene expression in the rainbow trout (Oncorhynchus mykiss) retina and hypothalamus. Juvenile fish were acclimated to a 12:12 light (L)-dark (D) cycle. Then, retina and hypothalamus were collected from animals kept under LD conditions or constant darkness (DD) for 24 h. Real-time quantitative reverse transcriptase–polymerase chain reaction (RT-PCR) assays were performed to quantify expression of the core circadian genes Clock1a, Bmal1, and Per1 as representative members of the circadian oscillator. All clock genes analyzed in the retina and hypothalamus showed circadian fluctuations. However, gene expression peaked in the rainbow trout hypothalamus with a 3-h (Clock1a and Bmal1) or 6-h (Per1) delay relative to that observed in the retina, the latter showing highest expression levels at zeitgeber times 9 (ZT9) for Clock1a and Bmal1, and at ZT21 for Per1. When exposed to DD, the rhythmic gene expression pattern was maintained for all genes in the rainbow trout retina, but only for Clock1a and Per1 in the hypothalamus. Bmal1 failed to cycle under DD, suggesting that hypothalamic clock function might depend on either several clock-gene isoforms or regulation from external inputs. Overall, these data indicate that representative molecular members of the core circadian clock are present in both the retina and hypothalamus of rainbow trout. (Author correspondence: jmmiguez@uvigo.es)

Separating the Contribution of Glucocorticoids and Wakefulness to the Molecular and Electrophysiological Correlates of Sleep Homeostasis

The sleep-deprivation-induced changes in delta power, an electroencephalographical correlate of sleep need, and brain transcriptome profiles have importantly contributed to current hypotheses on sleep function. Because sleep deprivation also induces stress, we here determined the contribution of the corticosterone component of the stress response to the electrophysiological and molecular markers of sleep need in mice. N/A SETTINGS: Mouse sleep facility. C57BL/6J, AKR/J, DBA/2J mice. Sleep deprivation, adrenalectomy (ADX). Sleep deprivation elevated corticosterone levels in 3 inbred strains, but this increase was larger in DBA/2J mice; i.e., the strain for which the rebound in delta power after sleep deprivation failed to reach significance. Elimination of the sleep-deprivation-associated corticosterone surge through ADX in DBA/2J mice did not, however, rescue the delta power rebound but did greatly reduce the number of transcripts affected by sleep deprivation. Genes no longer affected by sleep deprivation cover pathways previously implicated in sleep homeostasis, such as lipid, cholesterol (e.g., Ldlr, Hmgcs1, Dhcr7, -24, Fkbp5), energy and carbohydrate metabolism (e.g., Eno3, G6pc3, Mpdu1, Ugdh, Man1b1), protein biosynthesis (e.g., Sgk1, Alad, Fads3, Eif2c2, -3, Mat2a), and some circadian genes (Per1, -3), whereas others, such as Homer1a, remained unchanged. Moreover, several microRNAs were affected both by sleep deprivation and ADX. Our findings indicate that corticosterone contributes to the sleep-deprivation-induced changes in brain transcriptome that have been attributed to wakefulness per se. The study identified 78 transcripts that respond to sleep loss independent of corticosterone and time of day, among which genes involved in neuroprotection prominently feature, pointing to a molecular pathway directly relevant for sleep function.

Expression analysis of circadian genes in oocytes and preimplantation embryos of cattle and rabbits

We previously showed that circadian genes clock, bmal1, cry1, cry2, per1, and per2 are expressed and function as maternal mRNA regulating events in the oocytes and preimplantation embryos of mice. Recent evidence indicates however that either or both expression profiles of circadian genes in some tissues, and transcript sequences of circadian genes, differ to generate the physiological differences between diurnal and nocturnal species. We therefore investigated the expression profiles of circadian genes in oocytes and preimplantation embryos of species other than mice, namely cattle and rabbits, representing diurnal and nocturnal species, respectively, and determined the protein sequences of circadian genes in these species. Quantitative real-time PCR revealed that all circadian genes considered in this study were present in the oocytes and preimplantation embryos of both species, and the transcript amounts of clock, cry1 and per1 contained in oocytes were significantly higher than in preimplantation embryos of both species. The transcripts of clock, cry1, and per1 of cattle and rabbits were determined by primer walking, and functional domains in the estimated amino acid sequences were compared between cattle and rabbits and with those of humans and mice. The sequences of clock, cry1, and per1 in cattle and rabbits closely resembled those in mice (85–100% homologies), and no difference based on diurnality or nocturnality was observed. These findings suggest that circadian genes in the oocytes and preimplantation embryos of mammals fulfill the same functions across species as maternal mRNA.

Methylselenocysteine Resets the Rhythmic Expression of Circadian and Growth-Regulatory Genes Disrupted by Nitrosomethylurea In vivo

Epidemiologic and animal studies indicate that disruption of circadian rhythm increases breast cancer risk. Previously, we showed that methylselenocysteine reduced the incidence of N-nitroso-N-methylurea (NMU)-induced mammary carcinomas in Fischer 344 rats by 63%. Methylselenocysteine also increased the expression of Period 2 (Per2) and D-binding protein (DBP), providing evidence for a link between circadian rhythm and chemoprevention. Here, we report that NMU disrupted the expression of core circadian genes (Per1, Per2, Cry1, Cry2, and RevErbAalpha) and circadian-controlled genes, including melatonin receptor 1alpha (MTNR1A), estrogen receptors (ERalpha and ERbeta), and growth-regulatory genes (Trp53, p21, Gadd45alpha, and c-Myc) in mammary glands of Fischer 344 rats. By contrast, dietary methylselenocysteine (3 ppm selenium) given for 30 days, significantly enhanced the circadian expression of these genes (except for Cry1 and Cry2). The largest effect was on the levels of the Per2, MTNR1A, and ERbeta mRNAs, which showed 16.5-fold, 4.7-fold, and 9.5-fold increases in their rhythm-adjusted means, respectively, and 44.5-fold, 6.5-fold, and 9.7-fold increases in amplitude as compared with the control diet, respectively. Methylselenocysteine also shifted the peak expression times of these genes to Zeitgeber time 12 (ZT12; lights off). Methylselenocysteine also induced rhythmic expression of Trp53, p21, and Gadd45alpha mRNAs with peak levels at ZT12, when c-Myc expression was at its lowest level. However, methylselenocysteine had no significant effect on the circadian expression of these genes in liver. These results suggest that dietary methylselenocysteine counteracted the disruptive effect of NMU on circadian expression of genes essential to normal mammary cell growth and differentiation.

Зміни у рівні експресії циркадіальних генів Per1, Clock і BMal1 у печінці, легені, сім’янику, нирці та серці щурів за дії на організм наночасток срібла

Most of physiological and biochemical processes in living organisms have circa­dian character. Disturbance of the regulation of circadian processes leads to developing different pathological processes and cancer, in particular. In this work, we have studied the effect of silver nanoparticles on the expression of very important regulatory genes which control the circadian processes: Per1, Clock and BMal1. We have shown that the level of Per1 expression significantly increased in the liver and lung but decreased in the testes. The level of Clock mRNA expression also increased in liver and lung but decreased in kidney. The Clock mRNA expression in testis and heart was observed at late period of silver nanoparticles action. Thus, the expression of Per1, Clock and BMal1 mRNA significantly changed in tissue specific manner in rats treated with silver nanoparticles. Disturbance of these genes expression can destroy cellular signal pathways and leads to developing pathological processes. The results of our investigation clearly demonstrated that silver nanoparticles have significant effect on the regulatory mechanisms which control cell metabolism via circadian gene expression. Expression of circadian genes can be a sensitive test for the detection of silver nanoparticles action on the organisms. Keywords: Per1, Clock, BMal1, silver nanoparticles, liver, lung, testes, rats.

Regulation of αENaC Expression by the Circadian Clock Protein Per1

The mineralocorticoid aldosterone is a major regulator of sodium transport in target epithelia and an important determinant of systemic blood pressure. We have previously shown that knockdown of the circadian rhythm gene Per1 prevented the aldosterone‐mediated induction of the alpha subunit of the epithelial sodium channel (αENaC) in mouse inner medullary collecting duct cells. We have extended this observation to the outer medullary and cortical collecting duct using additional in vitro cell models. Interestingly, αENaC expression was attenuated following Per1 knockdown even in the absence of aldosterone, indicating that Per1 contributes to the basal regulation of αENaC expression. Analysis of heterogeneous nuclear RNA and αENaC promoter luciferase assays showed that this regulation occurred at the level of transcription. Most importantly, expression of αENaC was attenuated in Per1 deficient mice. Evaluation of αENaC mRNA levels over a 24 hour period revealed a circadian pattern of expression that was altered in mice lacking Per1. These data demonstrate a novel and functional role for the circadian clock protein Per1 in the regulation of the renal epithelial sodium channel. Supported by NIDDK, AHA.

Autism-associated SNPs in the clock genes npas2, per1 and the homeobox gene en2 alter DNA sequences that show characteristics of microRNA genes.

AbstractIntronic single nucleotide polymorphisms (SNPs) in the clock genes npas2 and per1 and the homeobox gene en2 are reported to be associated with autism. This bioinformatics analysis of the intronic regions which contain the autism-associated SNPs rs1861972 and rs1861973 in en2, rs1811399 in npas2, and rs885747 in per1, shows that these regions encode RNA transcripts with predicted structural characteristics of microRNAs. These microRNA-like structures are disrupted in silico by the presence of the autism enriched alleles of rs1861972, rs1861973, rs1811399 and rs885747 specifically, as compared with the minor alleles of these SNPs. The predicted gene targets of these microRNA-like structures include genes reported to be implicated in autism (gabrb3, shank3) and genes causative of diseases co-morbid with autism (mecp2 and rai1). The inheritance of the AC haplotype of rs1861972 - rs1861973 in en2, the C allele of rs1811399 in npas2, and the C allele of rs1234747 in per1 may contribute to the causes of autism by affecting microRNA genes that are co-expressed along with the homeobox gene en2 and the circadian genes npas2 and per1.

Chemopreventive Doses of Methylselenocysteine Alter Circadian Rhythm in Rat Mammary Tissue

It is known that organic forms of selenium inhibit chemically induced rat mammary carcinogenesis, although the molecular basis remains to be elucidated. To identify signaling pathways involved in carcinogenesis that are also modulated by methylselenocysteine, we compared the global gene expression profiles in mammary tissues from pubescent female rats maintained on a selenium-supplemented (3 ppm) diet with those on a standardized diet after N-nitroso-N-methylurea. Whereas the selenium-enriched diet altered the steady-state levels of genes involved in various cellular functions, the most dramatic effect was the coordinated changes in the expression of multiple genes that regulate circadian rhythm. Normal mammary tissue of rats fed a standardized diet showed little circadian oscillation relative to liver tissue. By contrast, mammary tissue of rats maintained on the selenium-enriched diet showed a progressive, time-dependent increase in the expression of circadian gene Per2 and circadian-regulated transcription factor DBP. Our results further showed that the expression of Per2 and DBP mRNAs was significantly decreased in mammary tumors arising in rats on the selenium-enriched diet, but not in tumors of rats on the control diet, suggesting that selenium-induced elevation in the expression of circadian genes was incompatible with mammary carcinogenesis. Given the previously reported role of Per2 as a tumor suppressor, these observations suggest that Per2 is an important target of methylselenocysteine during chemoprevention in N-nitroso-N-methylurea-induced rat mammary carcinogenesis, and for the first time provide a link between chemoprevention and circadian rhythm.