Association Study of a Variable-Number Tandem Repeat Polymorphism in the Clock Gene PERIOD3 and Chronotype in Norwegian University Students

Circadian rhythms are endogenously generated cycles involving physiological parameters, such as core body temperature, hormone levels, blood pressure, sleep, and metabolism, with a period length of around 24?h. The circadian clock in mammals is regulated by a set of clock genes that are functionally linked together, and polymorphisms in clock genes could be associated with differences in circadian rhythms. A variable-number tandem repeat (VNTR) in the human clock gene PERIOD3 (PER3) has been suggested to correlate with a morning (lark) versus evening (owl) chronotype as well as with the circadian rhythm sleep disorder “delayed sleep phase disorder” (DSPD). The authors examined 432 healthy Norwegian university students in search of further support for an association between the PER3 polymorphism and diurnal preference. The Horne-Östberg Morningness-Eveningness Questionnaire (MEQ) and Preferences Scale (PS) were used to evaluate subjective chronotype. DNA samples were genotyped with respect to the 4-repeat and 5-repeat alleles of the VNTR PER3 polymorphism, and the genotype distribution was 192 (4-4), 191 (4-5), and 49 (5-5). The authors estimated that the power to detect an association of the 4-allele with preference for morningness or eveningness was 75%. The authors found no association between the PER3 clock gene and chronotype, indicating that the proposed role of PER3 needs further clarification. (Author correspondence: teresa.osland@med.uib.no)     

A variable-number taandem repeat polymorphism in PER3 is not associated with chronotype in a population with self-reported sleep problems

We examined whether a variable-number tandem repeat (VNTR) polymorphism in the circadian clock gene PER3 was associated with subjective ratings of sleep and diurnal preference in a Romanian population with high levels of self-reported sleep problems. Individuals, self-reporting to their GP for sleep disturbances, completed a battery of validated scales that assess the presence of insomnia, sleep quality and diurnal preference and had their PER3 VNTR genotype determined. We found no significant differences in chronotype, sleep quality or other psychometric measures according to PER3 VNTR and conclude that diurnal preference or self-reported sleep measures are not associated with PER3 genotype in this population.

     

Phenotypic effects of genetic variability in human clock genes on circadian and sleep parameters

Circadian rhythms and sleep are two separate but intimately related processes. Circadian rhythms are generated through the precisely controlled, cyclic expression of a number of genes designated clock genes. Genetic variability in these genes has been associated with a number of phenotypic differences in circadian as well as sleep parameters, both in mouse models and in humans. Diurnal preferences as determined by the selfreported Horne-Östberg (HÖ) questionnaire, has been associated with polymorphisms in the human genes CLOCK, PER1, PER2 and PER3. Circadian rhythm-related sleep disorders have also been associated with mutations and polymorphisms in clock genes, with the advanced type cosegrating in an autosomal dominant inheritance pattern with mutations in the genes PER2 and CSNK1D, and the delayed type associating without discernible Mendelian inheritance with polymorphisms in CLOCK and PER3. Several mouse models of clock gene null alleles have been demonstrated to have affected sleep homeostasis. Recent findings have shown that the variable number tandem polymorphism in PER3, previously linked to diurnal preference, has profound effects on sleep homeostasis and cognitive performance following sleep loss, confirming the close association between the processes of circadian rhythms and sleep at the genetic level.     

Clock genes associate with white matter integrity in depressed bipolar patients

Human genetic studies have implicated specific genes that constitute the molecular clock in the manifestation of bipolar disorder (BD). Among the clock genes involved in the control system of circadian rhythms, CLOCK 3111 T/C and Period3 (PER3) influence core psychopathological features of mood disorders, such as patterns of sleep, rest, and activity, diurnal preference, cognitive performances after sleep loss, age at the onset of the illness, and response to antidepressant treatment. Furthermore, several studies pointed out that bipolar symptomatology is associated with dysfunctions in white matter (WM) integrity, suggesting these structural alterations as a possible biomarker of the disorder. We hypothesise that CLOCK and PER3 polymorphisms could be potential factors affecting WM microstructure integrity in bipolar patients. The relationship between these clock genes and DTI measures of WM integrity in a sample of 140 (53 M; 87 F) patients affected by BD type I was studied. Tract-based spatial statistics analyses on DTI measures of WM integrity were performed for each clock gene polymorphism, between the genetic groups. We accounted for the effect of nuisance covariates known to influence WM microstructure: age, sex, lithium treatment, age at the onset of the illness, and the number of illness episodes. We found that compared to T homozygotes, CLOCK C carriers showed a widespread increase of the mean diffusivity in several WM tracts. Compared with PER3^5/5 homozygotes, PER3^4/4 homozygotes showed significantly increased radial diffusivity and reduced fractional anisotropy in several brain WM tracts. No significant difference was observed between heterozygotes and the other subgroups. Altogether, this pattern of results suggests WM disruption in CLOCK C carrier and in PER3⁴ homozygotes. Sleep promotes myelination and oligodendrocyte precursor cell proliferation and associates with higher expression of genes coding for phospholipid synthesis and myelination in oligodendrocytes. These clock genes play a pivotal role in maintaining circadian rhythms and the sleep-wake cycle. Thus, it may be suggested that CLOCK rs1801260*C and PER3^4/4 influence myelination processes by regulating sleep quality and quantity.     

Higher scores in the extraversion personality trait are associated with a functional polymorphism in the PER3 gene in healthy subjects

A polymorphism in the PER3 (period circadian clock 3) gene has been associated with neuropsychiatric disorders and endophenotypes. We evaluated the possible association of personality domains with the PER3 polymorphism in a sample of healthy subjects: 271 individuals were evaluated with the Big Five Inventory and genotyped for the PER3 Variable Number Tandem Repeat (VNTR) polymorphism. We found a significant association between the PER3 polymorphism and the extraversion personality trait (p = 0.0093). The 5/5 genotype carriers showed higher scores for extraversion. This is the first time that a significant association between the PER3 VNTR polymorphism and extraversion is reported.     

PER3 Polymorphism Predicts Cumulative Sleep Homeostatic but Not Neurobehavioral Changes to Chronic Partial Sleep Deprivation

Background

The variable number tandem repeat (VNTR) polymorphism 5-repeat allele of the circadian gene PERIOD3 (PER3^5/5) has been associated with cognitive decline at a specific circadian phase in response to a night of total sleep deprivation (TSD), relative to the 4-repeat allele (PER3^4/4). PER3^5/5 has also been related to higher sleep homeostasis, which is thought to underlie this cognitive vulnerability. To date, no study has used a candidate gene approach to investigate the response to chronic partial sleep deprivation (PSD), a condition distinct from TSD and one commonly experienced by millions of people on a daily and persistent basis. We evaluated whether the PER3 VNTR polymorphism contributed to cumulative neurobehavioral deficits and sleep homeostatic responses during PSD.

Methodology/Principal Findings

PER3^5/5 (n = 14), PER3^4/5 (n = 63) and PER3^4/4 (n = 52) healthy adults (aged 22–45 y) demonstrated large, but equivalent cumulative decreases in cognitive performance and physiological alertness, and cumulative increases in sleepiness across 5 nights of sleep restricted to 4 h per night. Such effects were accompanied by increasing daily inter-subject variability in all groups. The PER3 genotypes did not differ significantly at baseline in habitual sleep, physiological sleep structure, circadian phase, physiological sleepiness, cognitive performance, or subjective sleepiness, although during PSD, PER3^5/5 subjects had slightly but reliably elevated sleep homeostatic pressure as measured physiologically by EEG slow-wave energy in non-rapid eye movement sleep compared with PER3^4/4 subjects. PER3 genotypic and allelic frequencies did not differ significantly between Caucasians and African Americans.

Conclusions/Significance

The PER3 VNTR polymorphism was not associated with individual differences in neurobehavioral responses to PSD, although it was related to one marker of sleep homoeostatic response during PSD. The comparability of PER3 genotypes at baseline and their equivalent inter-individual vulnerability to sleep restriction indicate that PER3 does not contribute to the neurobehavioral effects of chronic sleep loss.     

Period3 VNTR polymorphism influences the time-of-day pain onset of acute myocardial infarction with ST elevation

It is well established that the incidence and infarct size in acute myocardial infarction (AMI) is subject to circadian variations. At the molecular level, circadian clocks in distinct cells, including cardiomyocytes, generate 24-h cycles of biochemical processes. Possible imbalance or impairment in the cell clock mechanism may alter the cardiac metabolism and function and increase the susceptibility of cardiovascular diseases. One of the key components of the human clock system PERIOD3 (PER3) has been recently demonstrated to affect circadian expression of various genes in different tissues, including the heart. The variable number tandem repeat (VNTR) polymorphism (rs57875989) in gene Period3 (Per3) is related to multiple phenotypic parameters, including diurnal preference, sleep homeostasis, infection and cancer. The aim of our study was to investigate the effect of this polymorphism in AMI with ST elevation (STEMI). The study subjects (314 patients of Caucasian origin with STEMI, and 332 healthy controls) were genotyped for Per3 VNTR polymorphism using an allele-specific polymerase chain reaction. A gender difference in circadian rhythmicity of pain onset was observed with significant circadian pattern in men. Furthermore, the Per3^5/5 variant carriers were associated with higher levels of interleukin-6, B-type natriuretic peptide and lower vitamin A levels. By using cosinor analysis we observed different circadian distribution patterns of AMI onset at the level of genotype and allelic frequencies. Genotypes with at least one 4-repeat allele (Per3^4/5 and Per3^4/4) (N?=?264) showed remarkable circadian activity in comparison with Per3^5/5 (N?=?50), especially in men. No significant differences in genotype and/or allele frequencies of Per3 VNTR polymorphism were observed when comparing STEMI cases and controls. Our results indicate that the Per3 VNTR may contribute to modulation of cardiac functions and interindividual differences in development and progression of myocardial infarction.     

PER3 polymorphism predicts sleep structure and waking performance

Circadian rhythmicity and sleep homeostasis interact to regulate sleep-wake cycles [1-4], but the genetic basis of individual differences in sleep-wake regulation remains largely unknown [5]. PERIOD genes are thought to contribute to individual differences in sleep timing by affecting circadian rhythmicity [6], but not sleep homeostasis [7, 8]. We quantified the contribution of a variable-number tandem-repeat polymorphism in the coding region of the circadian clock gene PERIOD3 (PER3) [9, 10] to sleep-wake regulation in a prospective study, in which 24 healthy participants were selected only on the basis of their PER3 genotype. Homozygosity for the longer allele (PER3(5/5)) had a considerable effect on sleep structure, including several markers of sleep homeostasis: slow-wave sleep (SWS) and electroencephalogram (EEG) slow-wave activity in non-rapid eye movement (non-REM) sleep and theta and alpha activity during wakefulness and REM sleep were all increased in PER3(5/5) compared to PER3(4/4) individuals. In addition, the decrement of cognitive performance in response to sleep loss was significantly greater in the PER3(5/5) individuals. By contrast, the circadian rhythms of melatonin, cortisol, and peripheral PER3 mRNA expression were not affected. The data show that this polymorphism in PER3 predicts individual differences in the sleep-loss-induced decrement in performance and that this differential susceptibility may be mediated by its effects on sleep homeostasis.     

Sleep, diurnal preference, health, and psychological well-being: a prospective single-allelic-variation study

Individual differences in sleep and diurnal preference associate with physical and mental health characteristics, but few genetic determinants of these differences have been identified. A variable number tandem repeat (VNTR) polymorphism in the PERIOD3 (PER3) gene (rs57875989) has been reported to associate with diurnal preference, i.e., preferred timing of waking and sleep. Here, the authors investigate in a prospective single-candidate genetic variant study whether allelic variation for this polymorphism associates also with reported actual sleep timing and sleep duration, as well as psychological and health measures. Six hundred and seventy-five subjects, aged 20 to 35 yrs, completed questionnaires to assess sleep and psychological and health characteristics and were genotyped for the PER3 VNTR. Homozygosity for the longer allele (PER3(5/5)) of the VNTR was associated with increased morning preference, earlier wake time and bedtime, and reduced daytime sleepiness. Separate analyses of work and rest days demonstrated that the increase in time in bed during rest days was greatest in PER3(5/5) homozygotes. PER3 genotype modified the effects of sleep timing and duration on fluid intelligence and body mass index. Genotype was not associated with physical or psychological characteristics as assessed by the SF-36 Health Questionnaire, the General Health Questionnaire, the Big Five Inventory, the Behavioral Inhibition System-Behavioral Activation System scales, and the Positive and Negative Affect Scale, even though these measures varied significantly with diurnal preference as assessed by the Morningness-Eveningness Questionnaire. Whereas diurnal preference also predicts mental health and psychological characteristics, as well as sleep timing, the PER3 VNTR specifically affects measures of sleep timing and may also modify the effects of sleep on health outcome measures.     

PER3 polymorphism and cardiac autonomic control: effects of sleep debt and circadian phase

A variable number tandem repeat polymorphism in the coding region of the circadian clock PERIOD3 (PER3) gene has been shown to affect sleep. Because circadian rhythms and sleep are known to modulate sympathovagal balance, we investigated whether homozygosity for this PER3 polymorphism is associated with changes in autonomic nervous system (ANS) activity during sleep and wakefulness at baseline and after sleep deprivation. Twenty-two healthy participants were selected according to their PER3 genotype. ANS activity, evaluated by heart rate (HR) and HR variability (HRV) indexes, was quantified during baseline sleep, a 40-h period of wakefulness, and recovery sleep. Sleep deprivation induced an increase in slow-wave sleep (SWS), a decrease in the global variability, and an unbalance of the ANS with a loss of parasympathetic predominance and an increase in sympathetic activity. Individuals homozygous for the longer allele (PER3(5/5)) had more SWS, an elevated sympathetic predominance, and a reduction of parasympathetic activity compared with PER3(4/4), in particular during baseline sleep. The effects of genotype were strongest during non-rapid eye movement (NREM) sleep and absent or much smaller during REM sleep. The NREM-REM cycle-dependent modulation of the low frequency-to-(low frequency + high frequency) ratio was diminished in PER3(5/5) individuals. Circadian phase modulated HR and HRV, but no interaction with genotype was observed. In conclusion, the PER3 polymorphism affects the sympathovagal balance in cardiac control in NREM sleep similar to the effect of sleep deprivation.     

Chronotype of South African adults is affected by solar entrainment

Our daily lives are influenced by three different daily timers: the solar clock, our endogenous circadian clock and the societal clock. The way an individual’s endogenous clock synchronises to the solar clock, through either advances or delays relative to sunrise and sunset, results in a phenomenon known as diurnal preference or chronotype. South Africa uses just one time zone, but in the most easterly regions of the country, the sun rises and sets up to an hour earlier than in the most westerly regions throughout the year. It was hypothesised first that South Africans living in the east of the country may have a greater preference for mornings (more morning chronotypes) than those living in the west; and second, that this difference would not be due to genetic differences in the populations, particularly a genetic polymorphism previously shown to influence chronotype. Here, we describe and compare the distribution of chorotype and PERIOD3 variable number tandem repeat (VNTR) polymorphism frequency in eastern (n = 129) and western (n = 175) sample populations. Using the Horne–Östberg Morningness, Eveningness Questionnaire we found that there was a significantly higher proportion of morning-types in the eastern population (56.6%) than in the western population (39.4%), and there were higher proportions of neither-types and evening-types in the western population (51.4% and 9.1%, respectively) than in the eastern population (37.2% and 6.2%, respectively) (p = 0.009). There were no significant differences in distribution of the PER3 genotype (p = 0.895) and allele (p = 0.636) frequencies. Although previous studies have shown associations between chronotype and PER3 VNTR genotypes, no significant associations were observed in either the eastern (p = 0.695) or the western (p = 0.630) populations. These findings indicate that, in South African populations, longitude influences chronotype independently of PER3 genotype. The impacts of the differences in chronotype whilst maintaining the same societal temporal organisation in the eastern and western regions were not assessed.     

Association between Specific Diurnal Preference Questionnaire Items and PER3 VNTR Genotype

Although there are significant intra‐individual differences in self‐reported diurnal preference, as measured by validated questionnaires, the relative contribution of exogenous and endogenous factors to self‐reported diurnal preference largely remains to be investigated. The present study examined which items from the Horne‐Östberg (HÖ) questionnaire of diurnal preference were better at predicting genotypes in the variable number tandem polymorphism (VNTR) in the coding region of the gene PER3. This polymorphism has previously been reported to associate with diurnal preference, sleep parameters, and cognitive performance markers following sleep deprivation. Participants (n=240, selected from a previously studied population) had completed the HÖ questionnaire and provided a DNA sample, which was genotyped with regard to the PER3 VNTR. A multinomial logistic regression showed that four items significantly increased prediction accuracy between the two homozygotic genotypes, with homozygotes for the longer variant of the gene (PER3^5/5) associated with answers indicating a stronger morning preference than those chosen by homozygotes for the shorter variant (PER3^4/4). Only one item, the question of whether the respondent required an alarm clock, discriminated between all three genotypes. Moreover, when the items were divided into those with the strongest versus the weakest genetic association, there was a significant relationship between age and the questions not predicting genotype, but not between age and genotype‐predictive questions. This may explain previous findings regarding age‐related differences in self‐reported diurnal preference. These findings could facilitate the future development of diurnal preference scales especially tailored to the study of specific biological parameters.     

Sleep,Diurnal Preference,Health, and Psychological Well-being: A Prospective Single-Allelic-Variation Study

Individual differences in sleep and diurnal preference associate with physical and mental health characteristics, but few genetic determinants of these differences have been identified. A variable number tandem repeat (VNTR) polymorphism in the PERIOD3 (PER3) gene (rs57875989) has been reported to associate with diurnal preference, i.e., preferred timing of waking and sleep. Here, the authors investigate in a prospective single-candidate genetic variant study whether allelic variation for this polymorphism associates also with reported actual sleep timing and sleep duration, as well as psychological and health measures. Six hundred and seventy-five subjects, aged 20 to 35 yrs, completed questionnaires to assess sleep and psychological and health characteristics and were genotyped for the PER3 VNTR. Homozygosity for the longer allele (PER3^5/5) of the VNTR was associated with increased morning preference, earlier wake time and bedtime, and reduced daytime sleepiness. Separate analyses of work and rest days demonstrated that the increase in time in bed during rest days was greatest in PER3^5/5 homozygotes. PER3 genotype modified the effects of sleep timing and duration on fluid intelligence and body mass index. Genotype was not associated with physical or psychological characteristics as assessed by the SF-36 Health Questionnaire, the General Health Questionnaire, the Big Five Inventory, the Behavioral Inhibition System–Behavioral Activation System scales, and the Positive and Negative Affect Scale, even though these measures varied significantly with diurnal preference as assessed by the Morningness-Eveningness Questionnaire. Whereas diurnal preference also predicts mental health and psychological characteristics, as well as sleep timing, the PER3 VNTR specifically affects measures of sleep timing and may also modify the effects of sleep on health outcome measures. (Author correspondence: a.lazar@surrey.ac.uk)     

Effect of sleep deprivation on rhythms of clock gene expression and melatonin in humans

This study investigated the impact of sleep deprivation on the human circadian system. Plasma melatonin and cortisol levels and leukocyte expression levels of 12 genes were examined over 48?h (sleep vs. no-sleep nights) in 12 young males (mean?±?SD: 23?±?5 yrs). During one night of total sleep deprivation, BMAL1 expression was suppressed, the heat shock gene HSPA1B expression was induced, and the amplitude of the melatonin rhythm increased, whereas other high-amplitude clock gene rhythms (e.g., PER1-3, REV-ERBα) remained unaffected. These data suggest that the core clock mechanism in peripheral oscillators is compromised during acute sleep deprivation.     

Chronotype and PERIOD3 Variable Number Tandem Repeat Polymorphism in Individual Sports Athletes

A link between diurnal preference and a variable number tandem-repeat (VNTR) polymorphism in the PERIOD3 gene (PER3) has been demonstrated: the longer PER3(5) and shorter PER3(4) alleles with preferences for mornings and evenings, respectively. As many competitive events in South Africa for individual athletes are scheduled for the early mornings, we hypothesized that this might favor those athletes with a preference for morning activities. Self-selected white, male cyclists (CYC, n = 125), runners (RUN, n = 120) and Ironman triathletes (IM, n = 287) of European descent were compared with a control population of active, non-competitive individuals (CON, n = 96). The chronotypes of all CYC, RUN and CON participants and a sub-sample of the IM group (n = 49) were assessed using the Horne-?stberg Morningness-Eveningness Questionnaire, and the PER3 VNTR genotype for each participant was determined. The athlete groups contained more morning-type individuals than the CON group (CYC: 72%, n = 90; RUN: 67%, n = 80; IM: 59%, n = 29; CON: 41%, n = 39; p < .001). The prevalence of the PER3(5) allele was greater in the athlete groups (CYC: 61%, n = 152; RUN: 58%, n = 132; IM: 56%, n = 324; CON: 38%, n = 76; p < .001), and more athletes were genotyped as PER3(5/5) than CON individuals (CYC: 41%, n = 51; RUN: 23%, n = 26; IM: 28%, n = 81, CON: 9%, n = 8; p < .001). A strong relationship between chronotype and PER3 VNTR genotype was observed (p < .001). Finally, the time of day at which the athletes preferred to train was related to their chronotype (p < .001). This is the first study of its kind in a South African sporting population, and the results have not yet been replicated. These data suggest that white males of European descent participating in individual endurance sports in South Africa are more likely to be morning types. Furthermore, the PER3 VNTR may be one of the factors contributing to this observation. (Author correspondence: laura.roden@uct.ac.za ).     

Chronotype distribution in professional rugby players: Evidence for the environment hypothesis?

Individual sport athletes have been shown to comprise unusually high proportions of morning-types (MTs) coupled with a higher prevalence of the morningness-associated PERIOD3 variable number tandem repeat (VNTR) allele, PER3⁵. The degree to which type of sport selected is influenced by either chronotype or genotype, or the extent to which sporting environment contributes to chronotype is unclear. The aim of this study was to assess chronotype and PER3 VNTR polymorphism frequencies in team sport players and non-athletic controls. South African male Super Rugby players (RUG, n = 120) and a control population of males with habitually low levels of physical activity (defined as exercise no more than twice a week; CON, n = 117) took part in this study. Participants completed the Horne–Östberg morningness–eveningness questionnaire to determine chronotype and donated buccal cell or blood samples from which PER3 VNTR genotype was established. There were more MTs in the RUG (47%) than CON group (23%, p < 0.001), more evening-types in the CON group (18%) compared to the RUG group (3%, p < 0.001), but no differences in PER3 VNTR genotype (p = 0.619) or allele (p = 0.758) frequencies. In both groups, more people carried the PER3⁴ allele (RUG: 63%, CON: 62%). Chronotype was associated with genotype in the CON (p = 0.004) but not the RUG group (p = 0.895). Unlike the individual sport endurance athletes previously studied in whom the PER3⁵ allele predominated, the PER3 VNTR genotype distribution in these team sport players was similar to that of the general population. We hypothesise that the absence of any chronotype–genotype relationship in these rugby players is because their diurnal preference is shifted towards morningness through habitual athletic behaviour.     

Correlation among clock gene expression rhythms,sleep quality,and meal conditions in delayed sleep-wake phase disorder and night eating syndrome

Clock genes that comprise the circadian clock system control various physiological functions. Delayed sleep-wake phase disorder (DSWPD) and night eating syndrome (NES) are characterized by delayed sleep and meal timing, respectively. We estimated that clock gene expression rhythms in DSWPD patients may be delayed in comparison with the healthy subjects due to delayed melatonin secretion rhythms, producing eveningness chronotype in these individuals. However, it was difficult to estimate which clock gene expression rhythms were delayed or not in NES patients, because previous studies revealed that melatonin secretion rhythm was a little delayed compared with healthy individuals and that chronotype of NES patients depended on the individuals. Therefore, we examined expression rhythms of clock genes such as Period3 (Per3), nuclear receptor subfamily 1, group D, member 1 (Nr1d1) and Nr1d2 in these patients. Further, we expected sleep and meal patterns in DSWPD and NES patients may be more diverse than patterns observed in healthy subjects, and thus analyzed relationships among clock gene expression rhythms, sleep quality, sleep midpoint time, and meal times. We enrolled healthy male participants along with DSWPD and NES male patients, and asked all participants to answer questionnaires and to keep diaries to record information on their sleep and meals. Further, we asked them to collect 5–10 beard follicle samples, 6 times every 4 h. We measured clock gene expression rhythms using total RNA extracted from beard follicle cells. Peak time of clock gene expression in the NES group showed more diversity than the other groups, and that in the DSWPD group was delayed compared with the control group. In addition, the peak time of clock gene expression was negatively correlated with sleep quality and positively correlated with meal time after a long fast. Amplitudes of clock gene expression, especially Per3, positively responded to better mental and physical conditions as well as with better sleep quality. Results of this study suggest that peak times of clock gene expression in NES patients depended on the individuals; some patients with NES showed similar clock gene expression rhythm to healthy subjects, and other patients with NES showed similar to DSWPD patients. Moreover, this study suggests that meal time after a long fast may influence more determination in clock gene expression rhythms than the time of breakfast. Therefore, this study also indicates that Per3 clock gene may be one of the parameters that will help us understand sleep and meal rhythm disturbances.     

Evolutionary History of the PER3 Variable Number of Tandem Repeats (VNTR): Idiosyncratic Aspect of Primate Molecular Circadian Clock

The PER3 gene is one of the clock genes, which function in the core mammalian molecular circadian system. A variable number of tandem repeats (VNTR) locus in the 18th exon of this gene has been strongly associated to circadian rhythm phenotypes and sleep organization in humans, but it has not been identified in other mammals except primates. To better understand the evolution and the placement of the PER3 VNTR in a phylogenetical context, the present study enlarges the investigation about the presence and the structure of this variable region in a large sample of primate species and other mammals. The analysis of the results has revealed that the PER3 VNTR occurs exclusively in simiiforme primates and that the number of copies of the primitive unit ranges from 2 to 11 across different primate species. Two transposable elements surrounding the 18th exon of PER3 were found in primates with published genome sequences, including the tarsiiforme Tarsius syrichta, which lacks the VNTR. These results suggest that this VNTR may have evolved in a common ancestor of the simiiforme branch and that the evolutionary copy number differentiation of this VNTR may be associated with primate simiiformes sleep and circadian phenotype patterns.     

Rhythmic expression of circadian clock genes in human leukocytes and beard hair follicle cells

Evaluating individual circadian rhythm traits is crucial for understanding the human biological clock system. The present study reports characterization of physiological and molecular parameters in 13 healthy male subjects under a constant routine condition, where interfering factors were kept to minimum. We measured hormonal secretion levels and examined temporal expression profiles of circadian clock genes in peripheral leukocytes and beard hair follicle cells. All 13 subjects had prominent daily rhythms in melatonin and cortisol secretion. Significant circadian rhythmicity was found for PER1 in 9 subjects, PER2 in 3 subjects, PER3 in all 13 subjects, and BMAL1 in 8 subjects in leukocytes. Additionally, significant circadian rhythmicity was found for PER1 in 5 of 8 subjects tested, PER2 in 2 subjects, PER3 in 6 subjects, and BMAL1 in 3 subjects in beard hair follicle cells. The phase of PER1 and PER3 rhythms in leukocytes correlated significantly with that of physiological rhythms. Our results demonstrate that leukocytes and beard hair follicle cells possess an endogenous circadian clock and suggest that PER1 and PER3 expression would be appropriate biomarkers and hair follicle cells could be a useful tissue source for the evaluation of biological clock traits in individuals.