The moderating role of circadian gene polymorphisms in the relationship between sleep disturbance and circulating lymphocyte subsets in colorectal cancer patients.

AIM: We investigated the impact of sleep disturbance on immune status in colorectal cancer (CRC) patients with consideration of the moderating role of circadian clock gene polymorphisms. METHODS: A prospective longitudinal study design was used to collect information regarding sleep disturbance. Blood samples for immunologic assays were obtained the day before the first (baseline) and last cycles of 5-fluorouracil, leucovorin, and oxaliplatin (FOLFOX) chemotherapy. Clinical sleep disturbance was compared between the two-time points using the Pittsburgh Sleep Quality Index (PSQI) global score. We analysed single-nucleotide polymorphisms in rs2278749, rs3749474, rs2291738, rs17031614, and rs2287161. The dependent variables included changes in the percentages of CD4+, CD8+, CD19+, and CD16/56+ lymphocytes between the two-time points. The results were analysed using moderated regression analysis; the p-values were adjusted using the false discovery rate. RESULTS: Among the 104 patients, no significant dyadic associations were observed between changes in lymphocyte percentages and the PSQI global score. However, the moderated regression analysis revealed five significant associations (rs2287161 with CD8+, rs2278749 and rs2291738 with CD19+, and rs17031614 with CD4+ and CD16/56+ lymphocytes). The inclusion of each interaction resulted in a significant increase (5.7-10.7%) in the variance explained by changes in lymphocyte percentage. CONCLUSION: Patients with specific circadian gene allele types may be more susceptible to immune dysregulation when experiencing sleep disturbances. Considering that sleep disturbance is a modifiable factor that can impact immune regulation, it is essential to prioritise the management of sleep disturbances in CRC patients receiving FOLFOX chemotherapy.

The sex-dependent impact of PER2 polymorphism on sleep and activity in a novel mouse model of cranial-irradiation-induced hypersomnolence.

Background: Hypersomnolence is a common and disruptive side effect of cranial radiotherapy and is associated with fatigue and disturbances in mood and cognition in primary brain tumor (PBT) patients. The biological underpinnings of this effect are not understood. Our laboratory has previously found that the presence of a single nucleotide polymorphism (rs934945, G-E mutation) in the PERIOD2 (PER2) clock gene was associated with a decreased likelihood of fatigue in PBT patients. Here, we aim to understand the effects of PER2 polymorphism on radiation susceptibility within a murine model of cranial-irradiation-induced hypersomnolence (C-RIH). Methods: Male and female transgenic mice were generated using CRISPR-Cas9, replacing the endogenous mouse PER2:CRY1 binding domain with its human isoform with (hE1244 KI) or without the SNP rs934945 (hG1244 KI). Activity and sleep were monitored continuously 10 days before and after cranial irradiation (whole brain, 15Gy, single fraction). Behavioral assessments measuring anxiety, depression, and working memory were used to assess mood and cognitive changes 2 months postradiation. Results: During their active phase, hE1244 knock-ins (KIs) had less radiation-induced suppression of activity relative to hG1244 KIs and female hE1244 KIs saw a reduction of hypersomnolence over 10 days. hE1244 KIs displayed less anxiety behavior and were more ambulatory within all behavioral tests. Conclusions: The PER2 rs934945 polymorphism had long-lasting behavioral effects associated with radiation toxicity, particularly in sleep in females and the activity of all animals. Our findings shed light on biological mechanisms underlying C-RIH.

Ruscogenin timing administration mitigates cerebral ischemia-reperfusion injury through regulating circadian genes and activating Nrf2 pathway.

BACKGROUND: Ruscogenin (Rus), a steroidal sapogenin extracted from Ophiopogon japonicus (L. f.) Ker-Gawl., has the effect of alleviating cerebral ischemia-reperfusion injury (IRI), acute lung injury. At present, the chronopharmacological effects of Rus are still unknown. PURPOSE: This study explored the alleviating effect and mechanism of Rus timing administration on mice cerebral IRI. METHODS: The animals in different groups were administrated Rus (10 mg/kg) by gavage at four time points (23:00-01:00, 05:00-07:00, 11:00-13:00, 17:00-19:00) respectively for 3 days. On the 4th day, middle cerebral artery occlusion (MCAO) surgery was operated during 5:00-7:00. Behavioral tests were executed and the brain was collected for infarct volume, qPCR and immunoblot detection. The levels of tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), interleukin-1beta (IL-1ß) and inducible nitric oxide synthase (iNOS) were detected by qPCR. Glutathione (GSH), superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in serum and cerebral cortex were detected. The clock genes were tested by western blot. Based on these results, 17:00-19:00 was selected to administrate Rus for further mechanism study and Nrf2 blocker group was administrated all-trans-retinoic acid (ATRA) at 14:00 for 3 days. RESULTS: Administration of Rus reduced cerebral infarcted volume, ameliorated the behavior score and upregulated the mRNA and protein expression of Per1, Bmal1, Clock, Rev-erbα, transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), quinone oxidoreductase 1 (NQO1). Administration of Rus during 17:00-19:00 had better preventive effect than other three time points. Combined administration of ATRA blunted the preventive effect of Rus. CONCLUSION: The preventive effect of Rus is affected by the time of administration, which was regulated by Nrf2 pathway. Taken together, we provide solid evidence to suggest that different administration time point affect the effectiveness of Rus in alleviating IRI.

Circadian Gene Variants in Diseases.

The circadian rhythm is a self-sustaining 24 h cycle that regulates physiological processes within the body, including cycles of alertness and sleepiness. Cells have their own intrinsic clock, which consists of several proteins that regulate the circadian rhythm of each individual cell. The core of the molecular clock in human cells consists of four main circadian proteins that work in pairs. The CLOCK-BMAL1 heterodimer and the PER-CRY heterodimer each regulate the other pair's expression, forming a negative feedback loop. Several other proteins are involved in regulating the expression of the main circadian genes, and can therefore also influence the circadian rhythm of cells. This review focuses on the existing knowledge regarding circadian gene variants in both the main and secondary circadian genes, and their association with various diseases, such as tumors, metabolic diseases, cardiovascular diseases, and sleep disorders.

Rev-erbα agonist SR9009 protects against cerebral ischemic injury through mechanisms involving Nrf2 pathway.

Backgrounds: The circadian clock protein Rev-erbα is a crucial regulator of circadian rhythms that affects multiple molecular, cellular, and physiology pathways that control susceptibility, injury, and recovery in the neurological disorders. Emerging evidence suggest that Rev-erbα plays a key role in the inflammation and oxidative stress, two pivotal mechanisms in the pathogenesis, progression, and recovery process of ischemic stroke. However, it remains inconclusive whether Rev-erbα activation is protective against ischemic brain damage. Nuclear factor erythroid 2-related factor 2 (Nrf2) pathway, a master regulator of inflammatory and oxidative responses. Our study aimed to determine whether pharmacological activation of Rev-erbα by SR9009 protects against acute ischemic brain damage partly via Nrf2 pathway. Methods: Adult mice were pretreated with SR9009 or Nrf2 inhibitor all-trans-retinoic acid (ATRA) for 3 days prior to Sham or middle cerebral artery occlusion (MCAO) operation. After ischemia for 1 h and reperfusion for 24 h, the neurological function and cerebral infarction volume were determined, superoxide dismutase (SOD) activity, malondialdehyde (MDA) content and glutathione peroxidase (GSH-PX) activity in serum were detected by kit. The mRNA and/or protein level of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), inducible nitric oxide synthase (iNOS), Period (Per)1, Brain and muscle arnt-like1 (Bmal1), Circadian locomotor output cycles kaput (Clock), Rev-erbα, Nrf2, heme oxygenase-1 (HO-1) and quinone oxidoreductase 1 (NQO1) in cerebral cortex were detected by q-PCR and Western blot. Results: We confirmed that SR9009 activated Rev-erbα gene in the cerebral cortex under basal condition. At 24 h after reperfusion, SR9009 ameliorated acute neurological deficits, reduced infarct volume. Meanwhile, the inflammatory TNF-α, IL-1ß, iNOS and MDA content levels were significant decreased, SOD and GSH-PX activity were obviously increased, which were markedly blunted (or abolished) by ATRA. SR9009 enhanced the induction of Nrf2 and its downstream target genes HO-1 and NQO1 after ischemic insult. In addition, we found that SR9009 restored Rev-erbα, Bmal1, Clock, Per1 genes expression in the cerebral cortex under ischemic condition. Conclusion: Taken together, Rev-erbα activation by SR9009 protects against ischemic stroke damage, at least, partly through Nrf2 pathway.

Circadian Disruption and Breast Cancer Risk: Evidence from a Case-Control Study in China.

Studies had suggested an association between circadian disruptors (including night shift work, domestic light exposure at night, sleep duration, and circadian gene polymorphism) and breast cancer, while rare studies had been conducted in the Chinese population. This study was a case-control study conducted to explore the impact of circadian disruptors on the risk of breast cancer in China. Four hundred and sixty-four cases and 464 controls, admitted from the Department of Breast Surgery, Cancer Hospital, Chinese Academy of Medical Sciences, were included in this study. Adjusting age, BMI group, smoking, alcohol consumption, menopausal status, family history of breast cancer, duration of breastfeeding, age at menarche, number of pregnancies, age at first full-term pregnancy, use of estrogen and use of oral contraceptive, multivariate logistic regression analysis showed that the risk of breast cancer was higher in short sleep duration group (OR = 4.86, 95%CI: 1.73-17.33). Meanwhile, rs2292912 in CRY2, rs2253820 in PER1, rs2289591 in PER1 and rs3027188 in PER1 were positively associated with the risk of breast cancer. This study supported that the short duration of sleep and four SNPs in crucial circadian genes played a role in the development of breast cancer.

Advances in the Study of Circadian Genes in Non-Small Cell Lung Cancer.

Circadian genes regulate several physiological functions such as circadian rhythm and metabolism and participate in the cytogenesis and progression of various malignancies. The abnormal expression of these genes in non-small cell lung cancer (NSCLC) is closely related to the clinicopathological features of NSCLC and may promote or inhibit NSCLC progression. Circadian rhythm disorders and clock gene abnormalities may increase the risk of lung cancer in some populations. We collected 15 circadian genes in NSCLC, namely PER1, PER2, PER3, TIMELESS, Cry1, Cry2, CLOCK, BMAL1/ARNTL-1, ARNTL2, NPAS2, NR1D1(REV-ERB), DEC1, DEC2, RORα, and RORγ, and determined their relationships with the clinicopathological features of patients and the potential mechanisms promoting or inhibiting NSCLC progression. We also summarized the studies on circadian rhythm disorders and circadian genes associated with lung cancer risk. The present study aimed to provide theoretical support for the future exploration of new therapeutic targets and for the primary prevention of NSCLC from the perspective of circadian genes. Interpretation of circadian rhythms in lung cancer could guide further lung cancer mechanism research and drug development that could lead to more effective treatments and improve patient outcomes.

Relationship between circadian genes and memory impairment caused by sleep deprivation.

Background: Sleep deprivation (SD)-induced cognitive impairment is highly prevalent worldwide and has attracted widespread attention. The temporal and spatial oscillations of circadian genes are severely disturbed after SD, leading to a progressive loss of their physiological rhythms, which in turn affects memory function. However, there is a lack of research on the role of circadian genes and memory function after SD. Therefore, the present study aims to investigate the relationship between circadian genes and memory function and provide potential therapeutic insights into the mechanism of SD-induced memory impairment. Methods: Gene expression profiles of GSE33302 and GSE9442 from the Gene Expression Omnibus (GEO) were applied to identify differentially expressed genes (DEGs). Subsequently, both datasets were subjected to Gene Set Enrichment Analysis (GSEA) to determine the overall gene changes in the hippocampus and brain after SD. A Gene Oncology (GO) analysis and Protein-Protein Interaction (PPI) analysis were employed to explore the genes related to circadian rhythm, with their relationship and importance determined through a correlation analysis and a receiver operating characteristic curve (ROC), respectively. The water maze experiments detected behavioral changes related to memory function in SD rats. The expression of circadian genes in several critical organs such as the brain, heart, liver, and lungs and their correlation with memory function was investigated using several microarrays. Finally, changes in the hippocampal immune environment after SD were analyzed using the CIBERSORT in R software. Results: The quality of the two datasets was very good. After SD, changes were seen primarily in genes related to memory impairment and immune function. Genes related to circadian rhythm were highly correlated with engagement in muscle structure development and circadian rhythm. Seven circadian genes showed their potential therapeutic value in SD. Water maze experiments confirmed that SD exacerbates memory impairment-related behaviors, including prolonged escape latencies and reduced numbers of rats crossing the platform. The expression of circadian genes was verified, while some genes were also significant in the heart, liver, and lungs. All seven circadian genes were also associated with memory markers in SD. The contents of four immune cells in the hippocampal immune environment changed after SD. Seven circadian genes were related to multiple immune cells. Conclusions: In the present study, we found that SD leads to memory impairment accompanied by changes in circadian rhythm-related genes. Seven circadian genes play crucial roles in memory impairment after SD. Naïve B cells and follicular helper T cells are closely related to SD. These findings provide new insights into the treatment of memory impairment caused by SD.

PER2: a potential molecular marker for hematological malignancies.

Circadian rhythm is a periodic change of organism according to the law of external environment, which is manifested in metabolism, cell proliferation, physiology and behavior. In recent years, the role of circadian genes in the occurrence and progression of hematological malignancies have been continuously demonstrated. PER2 is the core component of the circadian rhythm playing an important role in regulating the circadian rhythm of the biological clock. This review summarizes the research progress of PER2 in hematological malignancies, especially leukemia, in order to better understand its role in hematological malignancies, and provide new ideas for clinical diagnosis and treatment.

NPAS2 Contributes to Liver Fibrosis by Direct Transcriptional Activation of Hes1 in Hepatic Stellate Cells.

Recently, emerging evidence shows that dysregulation of circadian genes is closely associated with liver fibrosis. However, how dysregulation of circadian genes promotes liver fibrosis is unknown. In this study, we show that neuronal PAS domain protein 2 (NPAS2), one of the core circadian molecules that has been shown to promote hepatocarcinoma cell proliferation, significantly contributed to liver fibrogenesis. NPAS2 is upregulated in hepatic stellate cells (HSCs) after fibrogenic injury, which subsequently contributes to the activation of HSCs. Mechanistically, NPAS2 plays a profibrotic role via direct transcriptional activation of hairy and enhancer of split 1 (Hes1), a critical transcriptor of Notch signaling for the fibrogenesis process, in HSCs. Our findings demonstrate that NPAS2 plays a critical role in liver fibrosis through direct transcriptional activation of Hes1, indicating that NPAS2 may serve as an important therapeutic target to reverse the progression of liver fibrosis.

Downregulation of the circadian Period family genes is positively correlated with poor head and neck squamous cell carcinoma prognosis.

We investigated the relationship between head and neck squamous cell carcinoma (HNSCC) and the mRNA and protein expression levels of the circadian genes of the Period (Per) family, Per1, Per2 and Per3. Tissue sections of HNSCC and normal head and neck tissues from two patient cohorts from two different hospitals were collected to assess the mRNA and protein expressions of the three Per family genes using real-time quantitative PCR (RT-PCR) and immunohistochemistry (IHC). The clinicopathological features and disease prognosis for the latter cohort were analyzed through IHC and statistical methods. Protein positive expression levels of the three Per family genes in HNSCC tissues was found to be approximately two times lower than that in normal tissues (p < .01). Moreover, patients with locally advanced HNSCC showed significantly greater downregulation of Per1, Per2 and Per3 mRNA expression levels as compared to patients with early-stage cancer (p < .05). Immunohistochemical examination of HNSCC patient tissues revealed a positive correlation between the Per family protein expression and the clinical tumor staging (p < .05). In addition, the Per protein-positive expression group showed higher 3-year survival rates [overall survival (OS) and progression-free survival (PFS)] as assessed by Kaplan-Meier plots and statistical analysis (p < .05). Our findings confirm the positive correlation between Per family gene expression and survival outcomes and support their role as prognostic markers for HNSCC.

Structural and mechanistic insights into the interaction of the circadian transcription factor BMAL1 with the KIX domain of the CREB-binding protein.

The mammalian CLOCK:BMAL1 transcription factor complex and its coactivators CREB-binding protein (CBP)/p300 and mixed-lineage leukemia 1 (MLL1) critically regulate circadian transcription and chromatin modification. Circadian oscillations are regulated by interactions of BMAL1's C-terminal transactivation domain (TAD) with the KIX domain of CBP/p300 (activating) and with the clock protein CRY1 (repressing) as well as by the BMAL1 G-region preceding the TAD. Circadian acetylation of Lys537 within the G-region enhances repressive BMAL1-TAD-CRY1 interactions. Here, we characterized the interaction of the CBP-KIX domain with BMAL1 proteins, including the BMAL1-TAD, parts of the G-region, and Lys537 Tethering the small compound 1-10 in the MLL-binding pocket of the CBP-KIX domain weakened BMAL1 binding, and MLL1-bound KIX did not form a ternary complex with BMAL1, indicating that the MLL-binding pocket is important for KIX-BMAL1 interactions. Small-angle X-ray scattering (SAXS) models of BMAL1 and BMAL1:KIX complexes revealed that the N-terminal BMAL1 G-region including Lys537 forms elongated extensions emerging from the bulkier BMAL1-TAD:KIX core complex. Fitting high-resolution KIX domain structures into the SAXS-derived envelopes suggested that the G-region emerges near the MLL-binding pocket, further supporting a role of this pocket in BMAL1 binding. Additionally, mutations in the second CREB-pKID/c-Myb-binding pocket of the KIX domain moderately impacted BMAL1 binding. The BMAL1(K537Q) mutation mimicking Lys537 acetylation, however, did not affect the KIX-binding affinity, in contrast to its enhancing effect on CRY1 binding. Our results significantly advance the mechanistic understanding of the protein interaction networks controlling CLOCK:BMAL1- and CBP-dependent gene regulation in the mammalian circadian clock.

Functional polymorphisms in circadian positive feedback loop genes predict postsurgical prognosis of gastric cancer.

BACKGROUND: Circadian positive feedback loop (CPFL) genes (CLOCK, BAML1, and NPAS2) have been implicated in cancer initiation and progression. The purpose of this study was to explore the effects of single-nucleotide polymorphisms (SNPs) in CPFL genes on prognosis of gastric cancer (GC) patients. METHODS: Nine functional SNPs from the three CPFL genes were genotyped in a cohort of 704 GC patients undergoing resection. Multivariate Cox regression model and Kaplan-Meier curve were used for prognosis analysis. RESULTS: Among the nine SNPs, rs11133399 in CLOCK, rs1044432 and rs2279284 in BAML1 were significantly associated with GC overall survival and recurrence-free survival. The unfavorable genotypes of these SNPs showed a cumulative effect on GC prognosis. Multivariate assessment model indicated that these SNPs, in conjunction with clinical variables, enhanced the power to predict GC prognosis. In addition, survival tree analysis revealed the genotype of rs11133399 as a primary risk factor contributing to the prognosis of GC patients. Functional assays showed that the G allele in rs11133399 significantly enhanced luciferase reporter activity than A allele. Immunohistochemical analysis further demonstrated that the genotype of rs11133399 was significantly associated with the expression level of CLOCK in GC tissues, suggesting that this SNP might affect the prognosis of GC through its influence on the expression of CLOCK gene. CONCLUSIONS: Our data indicate that SNPs in CPFL genes might contribute to the clinical outcome of GC through their impact on gene expression. Further studies are needed to elucidate its underlying molecular mechanisms.

Influence of the extracellular matrix on cell-intrinsic circadian clocks.

Cell-autonomous circadian clocks coordinate tissue homeostasis with a 24-hourly rhythm. The molecular circadian clock machinery controls tissue- and cell type-specific sets of rhythmic genes. Disruptions of clock mechanisms are linked to an increased risk of acquiring diseases, especially those associated with aging, metabolic dysfunction and cancer. Despite rapid advances in understanding the cyclic outputs of different tissue clocks, less is known about how the clocks adapt to their local niche within tissues. We have discovered that tissue stiffness regulates circadian clocks, and that this occurs in a cell-type-dependent manner. In this Review, we summarise new work linking the extracellular matrix with differential control of circadian clocks. We discuss how the changes in tissue structure and cellular microenvironment that occur throughout life may impact on the molecular control of circadian cycles. We also consider how altered clocks may have downstream impacts on the acquisition of diseases.

The correlation between PER3 rs2640908 polymorphism and colorectal Cancer in the Japanese population

Background: Colorectal cancer (CRC) is one of the most common cancers in Japan. Many factors influence this cancer, one of which is circadian rhythm disruption. Our research investigated the correlation between singlenucleotide polymorphisms (SNPs) in the Period 3 (PER3) (rs2640908), which is one of the circadian genes, and colorectal cancer in the Japanese population. Methods: The study participants consisted of 121 cases and 197 controls. DNA was extracted from participants' peripheral blood cells, and polymerase chain reaction­restriction fragment length polymorphism analysis (PCRRFLP) was performed to detect genotypes of PER3. Results: Participants with T/T genotype were at lower risk of developing colorectal cancer than participants with C/C genotype (adjusted ORs = 0.32 (95% CI: 0.15­0.63)). When stratified by gender and smoking status, T/T genotype were associated with a decreased susceptibility to cancer in males only (adjusted ORs: 0.23 (95% CI: 0.09­0.59)), T/T genotype were also associated with a decreased susceptibility to cancer among both smokers and non-smokers. Conclusions: A significant association was found between the T allele of PER3 polymorphism and a reduced risk of colorectal cancer, especially in males. Smoking status showed no association with the relationship between PER3 genotype and CRC carcinogenesis (AU)

Association between circadian gene CLOCK and cisplatin resistance in ovarian cancer cells: A preliminary study.

The present study aimed to observe the expression of circadian gene clock circadian regulator (CLOCK) in ovarian cancer cells and the effects of circadian gene CLOCK on cis-dichlorodiamine platinum (cisplatin) resistance in ovarian cancer cells. The expression of CLOCK mRNA and protein in cisplatin-sensitive A2780 and cisplatin-resistant CP70 cells were detected by quantitative polymerase chain reaction and western blot assay. Cisplatin-sensitive A2780 and cisplatin-resistant CP70 cells were treated with different concentrations of cisplatin for 48 h, and the expression of hCLOCK protein in the two types of cells was detected by western blot assay. RNA interference method was used to knock down the expression of CLOCK in cisplatin-resistant CP70 cells. Subsequently, the cisplatin-resistant CP70 cells were treated with cisplatin. The proliferation of cisplatin-resistant CP70 cells was observed following treatment with cisplatin. The expression of CLOCK mRNA was significantly higher in cisplatin-resistant CP70 cells (1.58±0.49) compared with cisplatin-sensitive A2780 cells (0.44±0.13) (P<0.01). Western blot assay results demonstrated that the expression of CLOCK protein was significantly greater in the cisplatin-resistant CP70 cells (1.47±0.34) compared with the cisplatin-sensitive A2780 cells (0.48±0.15) (P<0.01). Following the treatment of A2780 and CP70 cells with cisplatin, CLOCK protein expression increased with an increased concentration of cisplatin, in a dose-dependent manner (P<0.01). Following the knockdown of CLOCK in cisplatin-resistant CP70 cells by RNA interference, cisplatin treatment was able to significantly inhibit the proliferation of cells and induce apoptosis (P<0.01). The expression of circadian gene CLOCK in ovarian cancer cells was strongly associated with cisplatin resistance. The upregulation of circadian gene CLOCK in ovarian cancer cells may reduce its sensitivity to cisplatin treatment.

Epithelial and stromal circadian clocks are inversely regulated by their mechano-matrix environment.

The circadian clock is an autonomous molecular feedback loop inside almost every cell in the body. We have shown that the mammary epithelial circadian clock is regulated by the cellular microenvironment. Moreover, a stiff extracellular matrix dampens the oscillations of the epithelial molecular clock. Here, we extend this analysis to other tissues and cell types, and identify an inverse relationship between circadian clocks in epithelia and fibroblasts. Epithelial cells from mammary gland, lung and skin have significantly stronger oscillations of clock genes in soft 3D microenvironments, compared to stiff 2D environments. Fibroblasts isolated from the same tissues show the opposite response, exhibiting stronger oscillations and more prolonged rhythmicity in stiff microenvironments. RNA analysis identified that a subset of mammary epithelial clock genes, and their regulators, are upregulated in 3D microenvironments in soft compared to stiff gels. Furthermore, the same genes are inversely regulated in fibroblasts isolated from the same tissues. Thus, our data reveal for the first time an intrinsic difference in the regulation of circadian genes in epithelia and fibroblasts.

Temporal effects of human chorionic gonadotropin on expression of the circadian genes and steroidogenesis-related genes in human luteinized granulosa cells.

OBJECTIVE: It has been shown in animal models that circadian clock exists in corpora luteum which is essential for maintaining pregnancy. However, it is unknown whether circadian clock exists in corpora luteum and its relation with steroidogenesis in human ovary. STUDY DESIGN: Human luteinized granulosa cells from patients who underwent in vitro fertilization treatment were purified and cultured in vitro. Accumulation patterns of circadian gene and steroidogenesis-related gene mRNAs in human luteinized granulosa cells were observed during the 48 hours after treatment with human chorionic gonadotropin (hCG) by quantitative PCR. RESULTS: We found that the circadian genes CLOCK, PER2, and BMAL1 were expressed in cultured human luteinized granulosa cells. Among these genes, only expression of PER2 displayed oscillating patterns with a 16-h period in these cells after stimulation by hCG. Expression of CLOCK and BMAL1 did not show significant oscillating patterns. Expression of the steroidal acute regulatory protein (STAR) gene showed an oscillating pattern that was similar to that of PER2. Expression of CYP11A1, HSD3B2, and CYP19A1 increased significantly after hCG stimulation; however, none of these genes displayed significant oscillating patterns. CONCLUSIONS: Molecular circadian clock exists in human luteinized granulosa cells and may be related with steroidogenesis in human ovary.

Yin-yang actions of histone methylation regulatory complexes in the brain.

Dysregulation of histone methylation has emerged as a major driver of neurodevelopmental disorders including intellectual disabilities and autism spectrum disorders. Histone methyl writer and eraser enzymes generally act within multisubunit complexes rather than in isolation. However, it remains largely elusive how such complexes cooperate to achieve the precise spatiotemporal gene expression in the developing brain. Histone H3K4 methylation (H3K4me) is a chromatin signature associated with active gene-regulatory elements. We review a body of literature that supports a model in which the RAI1-containing H3K4me writer complex counterbalances the LSD1-containing H3K4me eraser complex to ensure normal brain development. This model predicts H3K4me as the nexus of previously unrelated neurodevelopmental disorders.

Role of circadian gene Clock during differentiation of mouse pluripotent stem cells.

Biological rhythms controlled by the circadian clock are absent in embryonic stem cells (ESCs). However, they start to develop during the differentiation of pluripotent ESCs to downstream cells. Conversely, biological rhythms in adult somatic cells disappear when they are reprogrammed into induced pluripotent stem cells (iPSCs). These studies indicated that the development of biological rhythms in ESCs might be closely associated with the maintenance and differentiation of ESCs. The core circadian gene Clock is essential for regulation of biological rhythms. Its role in the development of biological rhythms of ESCs is totally unknown. Here, we used CRISPR/CAS9-mediated genetic editing techniques, to completely knock out the Clock expression in mouse ESCs. By AP, teratoma formation, quantitative real-time PCR and Immunofluorescent staining, we did not find any difference between Clock knockout mESCs and wild type mESCs in morphology and pluripotent capability under the pluripotent state. In brief, these data indicated Clock did not influence the maintaining of pluripotent state. However, they exhibited decreased proliferation and increased apoptosis. Furthermore, the biological rhythms failed to develop in Clock knockout mESCs after spontaneous differentiation, which indicated that there was no compensational factor in most peripheral tissues as described in mice models before (DeBruyne et al., 2007b). After spontaneous differentiation, loss of CLOCK protein due to Clock gene silencing induced spontaneous differentiation of mESCs, indicating an exit from the pluripotent state, or its differentiating ability. Our findings indicate that the core circadian gene Clock may be essential during normal mESCs differentiation by regulating mESCs proliferation, apoptosis and activity.