The role of circadian clock in regulating cell functions: implications for diseases

Macrophages possess a robust circadian clock that plays a crucial role in driving their inflammatory and immune functions (Figure 3).66 About 8% of macrophage transcriptome including clock factors, inflammatory cytokines, and regulators of inflammatory cytokines such as pathogen recognition receptors and inflammatory signaling pathways oscillates in a circadian manner.67 A previous study identified 1403 genes, which account for 8.1% of the total, to be rhythmically expressed in a circadian manner.68 Another study involving transcriptome and proteome profiling demonstrates that only 15% of circadian proteome show corresponding oscillating mRNA patterns in murine bone marrow‐derived macrophages, suggesting that post‐transcriptional regulation plays a significant role in affecting the clock regulatory output in macrophages.69 This observation may be attributed to the abundance of proteins involved in degradation and translation. Macrophages exhibit unique circadian post‐transcriptional regulation due to their need for prompt response to a wide range of potential immunological stimuli.69

Clock genes, cytokines, and chemokines exhibit robust circadian rhythms in macrophages （Figure 3）.66, 70 Clock genes such as Rev‐erbα and Per2 display rhythmic expressions with high‐amplitude diurnal oscillations in peritoneal macrophages and central nervous system resident macrophages (microglia).67, 71 Tumor necrosis factor (TNF)‐α and interleukin (IL)‐6 secretion are observed in lipopolysaccharides (LPS)‐treated spleen macrophages with a peak phase around the transition time from light to dark, suggestive of a circadian clock‐regulated macrophage‐dependent cytokine secretion.67 Pro‐inflammatory cytokines show higher levels during the light phase in hippocampal microglia.72 Disruption or attenuation of core clock genes often disturbs macrophage circadian behaviors or functions. For example, circadian disruption induced by chronic jet lag modifies the rhythmic patterns of M1/M2 macrophages and cytokine levels in the spleen and tumor tissues.73 Deficiency of Rev‐erbα abolishes the circadian rhythmicity of IL‐6 in LPS‐induced macrophages and endotoxin‐treated mice.74Bmal1 deficiency decreases pro‐inflammatory cytokines (e.g., IL‐1β, TNF‐α, and IL‐6) and increases anti‐inflammatory cytokines (e.g., IL‐10) in microglial BV2 cells following LPS stimulation.75 In glioblastoma, deletion of Clock or its target gene OLFML3 (a microglia‐attracting chemokine) decreases intratumoral microglia infiltration in the tumor microenvironment and increases overall survival.76 Pathogen recognition receptors, a group of innate immune receptors, serve as upstream regulators of cytokines/chemokines such as IL‐1, TNF‐α, and IL‐6 through shared signaling modules including NF‐κB, activator protein‐1 (AP‐1), and MAPK,77 and regulation of NF‐κB, AP‐1, and MAPK by circadian clocks may contribute to the diurnal variations of cytokines/chemokines.78, 79, 80

Neutrophils are the predominant leukocyte subset in blood. The numbers, behaviors, and functions of neutrophils show apparent diurnal oscillations (Figure 3). The infiltration of neutrophils displays differential circadian rhythmicity in specific subsets and tissues.81 Under steady‐state conditions, neutrophil counts in the blood show circadian fluctuations.82, 83 In both humans and mice, peripheral neutrophil numbers decrease during the rest phase, whereas their numbers increase during the active phase.84 The levels of CD62⁻ CXC‐chemokine receptor (CXCR) 4⁺ neutrophils in circulation also follow a circadian rhythm with a nadir in the early active phase in mice. Consistently, blood neutrophil counts are lowest in the early morning in the UK Biobank participants.83, 85 This is because the release of young neutrophils and clearance of aged neutrophils from the periphery exhibit a time‐dependent manner.83, 85

Neutrophil homing to most naïve tissues such as lung and heart exhibits diurnal variations with a peak at night, whereas infiltration of neutrophils shows no circadian rhythms in the intestine, liver, and white adipose tissue.86 The ability of neutrophils to form neutrophil extracellular traps (NETs) also follows a circadian oscillation. In humans, the capacity of granule‐loading and NETs formation in neutrophils are higher at around 8:00 a.m. and gradually decrease later in the day.87 In mice, neutrophils express more enhanced levels of proteins related to granules, NETs generation, and cell migration in the nighttime.88 Specific deletion of Bmal1 abolishes daily variations in granule contents and NETs formation.89 Some markers such as CD11a, intercellular adhesion molecule‐1, L‐selectin, and the chemokine receptor CXCR4 in neutrophils exhibit circadian oscillations in humans, while P‐selectin glycoprotein ligand‐1 (PSGL‐1), L‐selectin, CD11a, and CD29 display rhythmic expression in mice (Figure 3).81 Antagonistic chemokine signaling drives neutrophil release from bone marrow through two main receptors CXCR4 and CXCR2. BMAL1 coordinates diurnal changes in the transcriptional and migratory properties of circulating neutrophils by regulating chemokine CXCL2 and chemokine receptor CXCR2 (Figure 3).90 Genetic disruption of the circadian clock has an impact on neutrophil trafficking. CXCL12/CXCR4 axis controlled by the central clock underlies the circadian variations of neutrophil numbers in blood (Figure 3). Genetic deletion of Cxcr4 in myeloid cells leads to a blunted oscillation of neutrophil infiltration into the blood.91 Downregulation of CXCL12 in the daytime stimulates the egress of neutrophils from the bone marrow into the blood.92 In addition, ablation of Bmal1 in myeloid cells leads to a lack of rhythmic donor neutrophil migration into the spleen, which is attributed to a reduced PSGL‐1 (an adhesion molecule involved in immune cell trafficking) expression.93

DCs play a pivotal role in the adaptive immune system, orchestrating antigen phagocytosis, processing, and presentation to naïve T cells, thereby initiating immune responses in pathological settings. The intricate orchestration of this process involves the expression of core clock genes (i.e., Per1, Per2, Bmal1, and Clock) and CCGs within DCs, which exhibit inherent circadian rhythmicity (Figure 3).94 Synchronized DCs demonstrate robust oscillations in the expressions of Bmal1, Rev‐erbα, and Per2.

The circadian rhythm regulates the differentiation, behaviors, and functions of DCs. First, loss of Rev‐erbs increases the expressions of surface molecules and genes (i.e., major histocompatibility complex II [MHCII] and CD86) that are important for DC maturation and immune responses. Besides, Rev‐erbα deficiency shows no effects on CD11b⁺CD11c⁺ DC populations, whereas Rev‐erbβ deficiency increases the percentage of CD11b⁺CD11c⁺ DC cells, suggesting a unique role of Rev‐erbβ in CD11b⁺CD11c⁺ DC development.95 In addition, the circadian rhythm of antigen processing in DCs is noticeably disrupted upon deletion of Bmal1. These findings collectively highlight the substantial contribution of molecular clock in governing the antigen‐processing capabilities of DCs.96 Furthermore, migration of DCs into lymph nodes also exhibits a circadian pattern, which peaks during the rest phase in mice (Figure 3).82 Circadian‐regulated functionality of DCs intricately shapes adaptive immune responses. For instance, intravenous administration of ovalbumin‐loaded DCs into mice in the daytime prominently augments the population of CD8+ T cells within the spleen, compared to nighttime administration.96

Biological clocks have emerged as crucial regulators of the trafficking DCs into lymph nodes. First, circadian clock genes are essential for maintaining the rhythmic migration of DCs into lymph nodes, involving both lymphatic endothelial cells (LECs) and DCs. Circadian expression of factors such as CC chemokine ligand 21 (CCL21), lymphatic vessel endothelial hyaluronan receptor 1, CD99, junctional adhesion molecule (JAM)‐A, and JAM‐C, which are associated with DC migration, is found to be dependent on the presence of functional BMAL1 in LECs. Notably, rhythmic expression of CCR7, a receptor for CCL21, is directly modulated by BMAL1 in DCs (Figure 3).97 BMAL1 also plays a significant role in DC antigen processing by influencing Ca²⁺ localization, mitochondrial dynamics, and metabolic processes, thereby impacting adaptive immune responses.96 Additionally, REV‐ERBα has been identified as a contributor to DC development, as it enhances the expression of MHCII and the co‐stimulatory receptor CD86, both of which serve as markers for DC maturation (Figure 3).95

Lymphocytes, as a prominent subset of immune cells responsible for immune recognition, exhibit significant circadian alterations in their numbers, including granulocytes and lymphocytes (Figure 4). T lymphocytes, a principal subset of lymphocytes, are identifiable through surface markers such as CD3, CD4, and CD8. These cells, upon activation, initiate a process of proliferation and differentiation, culminating in the secretion of cytokines, notably interferon (IFN)‐γ and IL‐2. Diverse subtypes of T cells, notably within the CD4+ and CD8+ subsets, display circadian rhythms. The number of antigen‐stimulated IFN‐γ+CD4+ T cells is higher during the nighttime, while their proportion significantly decreases during the daytime.98 CD4‐positive T cells exhibit pronounced circadian oscillations in various parameters, including clock gene expressions, cytokine release (IL‐2, IL‐4, IFN‐γ), and CD40L expression post stimulation (Figure 4).98 Natural regulatory T cells (T_reg), crucial in shaping adaptive immune responses, exhibit a clear circadian rhythm, with their highest levels occurring at night and lowest during the day.99 Additionally, molecular clocks within T cells play a role in regulating their trafficking behaviors, including homing and egress within lymph nodes.100 Circadian clocks also participate in the regulation of T‐cell differentiation. It has been established that molecular clocks, such as BMAL1, CLOCK, and REV‐ERBα, directly govern the differentiation of T helper 17 (T_H17) cells.101

The rhythmic oscillations of T‐cell numbers, reactivity, functions, and trafficking behaviors are reliant on biological clocks or circadian cues. To begin with, the circadian rhythm of T‐cell numbers is primarily affected by endogenous cortisol production, characterized by higher concentrations in the early morning and lower concentrations at midnight. This cortisol production is under the control of the circadian clock (Figure 4).102 Recruitment of T cells to lymph nodes is determined by the rhythmicity of T cells themselves as well as the microenvironment, which are regulated by CCR7–CCL21 axis. Additionally, oscillatory expression of sphingosine 1‐phosphate receptor 1 (S1pr1), controlled by BMAL1 and CLOCK, governs the rhythmic egress of T cells (Figure 4).101 Moreover, NF‐κB pathway influenced by the cell‐intrinsic clock has been identified as a potential mechanism mediating the rhythmicity of T‐cell immune responses (Figure 4).100 Furthermore, diurnal variations in T‐cell function are influenced by the circadian oscillation of DC function, which is regulated by the biological clock.96 Last, ablation of Clock has been found to diminish the capacity for T_H17‐cell differentiation and reduce T_H17‐cell frequencies in the gut through the regulation of retinoid‐related orphan receptor gamma t (Rorγt) transcription and Nfil3 expression (Figure 4).102

B cells, as a crucial component of the adaptive immune system, display diurnal variations in their numbers. In human blood, B‐cell numbers exhibit a significant peak during nighttime, followed by decreased levels during the daytime.103 Additionally, B‐cell counts in lymph nodes also display rhythmicity, with the highest levels during the early dark phase, which is closely associated with circadian behaviors such as homing and egress (Figure 4).101 Moreover, mRNA levels of canonical clock genes, including Bmal1, Clock, and Per2, exhibit oscillations in splenic B cells under the daily light–dark cycle and constant conditions.104 B‐cell receptor (BCR) signaling pathway plays a critical role in the activation, proliferation, and differentiation of B cells. Notably, loss of CRY, a circadian protein, leads to a significant activation of BCR‐proximal signaling pathways and affects B‐cell functions.105 Hence, circadian clock plays a vital role in modulating B cell numbers, development, and functions.

Regulation of B‐cell numbers, development, and functions relies on both circadian clocks and external cues. First, the diurnal rhythm in circulating B‐cell numbers in humans is influenced by the rhythmic plasma cortisol levels. This diurnal oscillation of B‐cell numbers coincides inversely with plasma cortisol levels.106 Second, the biological clock also plays a crucial role in modulating B‐cell numbers. Genetic deletion of clock proteins has been shown to impair B‐cell development and leads to a significant reduction in B‐cell populations in peripheral blood, spleen, and bone marrow.107 Additionally, loss of Bmal1 specifically in B cells disrupts the overall oscillation of B‐cell numbers in lymph nodes by regulating the diurnal expressions of key regulators such as Ccr7 and S1pr1, which are essential for lymphocyte trafficking (Figure 4).101 Moreover, CLOCK forms a complex with c‐Maf inducing protein (CMIP) to induce the degradation of CMIP, resulting in restricted transcription of the Il‐10 gene in B cells. This, in turn, suppresses the immune regulatory functions of B cells (Figure 4).28

The ILC family is stratified into two principal subtypes: cytotoxic and helper‐like ILCs. The cytotoxic subset encompasses natural killer (NK) cells, renowned for their inherent ability to target and eliminate malignant or pathogen‐infected cells. Conversely, the helper‐like ILCs are segregated into three primary groups: ILC1, ILC2, and ILC3. Each group exhibits distinct functional characteristics and plays a pivotal role in orchestrating immune responses.

NK cells, a crucial subset of lymphocytes distinguished by the absence of the CD3‐T cell receptor (TCR) complex, play a pivotal role in safeguarding the host against microbial infections and malignant transformations by means of discharging cytolytic granules or TNF‐α and triggering the Fas pathway.108 Recent studies have elucidated that within NK cells, the expression patterns of core circadian genes, including Bmal1, Clock, Per1, and Per2, display a distinct circadian rhythm. This rhythmic expression extends to cytolytic granules, notably granzyme B and perforin, as well as NK cell‐associated cytokines such as IFN‐γ and TNF‐α. These findings indicate a time‐dependent modulation of NK cells’ cytolytic activity, underpinned by the circadian oscillation of both gene expressions and effector functions (Figure 4).109 Additionally, circulating NK cell counts undergo a 24‐h oscillation in humans, displaying peak levels during the early morning and diminishing during the night (Figure 4).110 Nevertheless, the underlying mechanism responsible for the diurnal variations in NK cell numbers remains elusive.

Disruption of clock genes through genetic manipulation or exposure to environmental factors has provided valuable insights into the distinct roles of biological clocks in regulating the functions of NK cells. For instance, specific knockdown of Per2 using RNA interference techniques causes a significant decrease in the levels of granzyme B and perforin within the NK cell line RNK16.111 Additionally, loss of Per1 alters the rhythmic levels of perforin, granzyme B, and IFN‐γ released by splenic NK cells (Figure 4).112 Furthermore, chronic shift‐lag condition accelerates the aging process of NK cells, characterized by a reduced expression of Ly49 and impaired releases of granular CD107a and IFN‐γ.113 Repeated phase adjustments in the light–dark cycle similarly disrupt the rhythmic expressions of clock genes, cytolytic factors, and cytokines in NK cells, thereby impeding their time‐dependent cytolytic activities.114

In comparison to ILC1 and ILC2, ILC3 exhibits higher expressions of a range of clock genes, such as Bmal1, Clock, Nfil3, Dbp, Pers, Crys, and Rev‐erbs, which shows more distinct rhythmicity.23 The interplay between circadian clocks and ILC3 has garnered increasing attention in recent research. External cues like light exposure, dietary patterns, and microbiota composition, along with intrinsic circadian mechanisms, have been identified as modulators of the rhythmic expressions of clock genes (i.e., Clock and Rev‐erbα) in ILC3.23 Furthermore, circadian clock exerts a regulatory influence over the quantity, behavior, and functional attributes of ILC3. Notably, ablation of Bmal1 leads to a marked decrease in the population of ILC3 within the small intestine of mice.115 REV‐ERBα, a pivotal transcription factor for ILC3, significantly influences various ILC3 subsets.116 Perturbations in circadian rhythm have been observed to impact cytokine secretion in ILC3. For instance, Rev‐erbα deficiency substantially diminishes the numbers of NKp46+ ILC3 cells, reduces RORγt expression, and lowers IL‐22 production, while paradoxically elevating IL‐17 secretion.116 In ILC3, BMAL1 deficiency shows enhanced expressions of RORγt‐dependent target genes, including IL17a, IL17f, and IL22.115 Investigations into various types of ILCs, beyond ILC3, remain relatively sparse. However, notable advancements have been made in understanding the role of RORα in the differentiation of ILC2. These cells are integral to type II inflammation, allergic responses, and defense mechanisms against parasitic infections. The significance of RORα in the differentiation process of ILC2 underscores the critical functions of these cells in the immune response.61 Additionally, ILC2, which is under circadian regulation, play a vital role in the maintenance of eosinophil homeostasis.117

The circadian regulation of helper‐like ILCs is governed by a multitude of mechanisms. BMAL1 deficiency leads to a decrease in ILC3 counts in the small intestine, and an increased presence of ILC3s in the mesenteric lymph nodes attributed to loss of gut homing signals such as CCR9, α4β7 integrin, and CXCR4.23 Furthermore, gut microbiota and pro‐apoptotic proteins, such as Bim and Bax, are identified as key factors in controlling the hyper‐activation and loss of intestinal ILC3 in BMAL1‐deficient mice. The elimination of microbiota through antibiotic treatment partially mitigates the hyper‐activation of Bmal1‐deficient ILC3 and restores cellular homeostasis in the gut.115 However, regulatory mechanisms of helper‐like ILCs by circadian clocks have remained largely unexplored.

Hepatocytes, as the principal parenchymal cells in the liver, orchestrate a myriad of cellular processes including drug metabolism, drug detoxification, and immune cell activation, all critical for preserving liver homeostasis.118 The circadian clock mechanism exerts regulatory control over the diurnal rhythmicity of gene expressions and functional processes within hepatocytes. Perturbation of REV‐ERBs disrupts the natural diurnal rhythm and induces alterations in the metabolomic profiles observed across diverse liver cell categories.122 Furthermore, disruption of CLOCK function results in modified diurnal patterns of cytochrome P450 enzymes within hepatocytes, including but not limited to CYP2A4/5 and CYP2B10.123

The hepatocyte clock plays a vital role in sustaining rhythmic gene expression patterns in immune cells. Specifically, hepatocyte‐specific knockout of Rev‐erbs leads to marked changes in the rhythmic gene expression profiles of nonparenchymal liver cells, with macrophages being notably affected.124 Hepatocyte‐targeted Rev‐erbs knockdown disrupts the rhythmicity of genes linked to the vitamin D receptor signaling pathway, a pathway known to mitigate liver inflammation and steatosis. It also enhances the circadian rhythmicity of genes associated with inflammatory responses in macrophages.124, 125, 126 The interplay between hepatocytes and macrophages, facilitated by mediators including hepatocyte‐derived sterol regulatory‐element binding protein (SREBP) cleavage‐activating protein and a range of polypeptides, plays a critical role in the transmission of circadian rhythmic signals.124 In conclusion, circadian clocks are integral to the regulation of gene expressions and cellular functions in hepatocytes, underscoring their role in maintaining hepatic and systemic physiological balance.

Adipose tissue dynamically responds to the daily fluctuations in energy availability and demand, a crucial aspect of maintaining energy homeostasis. This adaptability is facilitated by a circadian modulation of signals, where adipose physiology is governed in a time‐of‐day dependent manner. This rhythmic regulation is primarily achieved through the transcriptional control of CCGs, which are regulated by adipocyte clocks.127 Adipocyte clocks orchestrate the timing of the local transcriptome, which includes critical genes involved in lipogenesis and lipolysis, such as Atgl, Hsl, caveolin 2, acyl‐CoA synthetase, phosphatidate phosphatase, Lpl, peroxisome proliferator‐activated receptor α/γ, coactivator 1β, and Srepb1α.127

The BMAL1/CLOCK heterodimer in adipocyte assumes a central role in the regulation of gene expressions, specifically activating the transcription of Atgl and Hsl genes by binding to E‐box promoter elements.128 Notably, adipocyte‐selective deletion of Rev‐erbα, under normal conditions, exhibits a relatively modest impact on the expressions of lipogenic genes. However, in the context of diet‐induced obesity, this deletion exerts a profound influence on metabolic health. It leads to the development of a metabolically advantageous phenotype characterized by diminished adipose tissue inflammation and distinct alterations in the dynamics of collagen fibrosis. These effects are attributed to the regulatory control of collagen genes.129 Furthermore, circadian clock exerts temporal control over adipogenic differentiation. BMAL1 transcriptionally regulates genes associated with the actin dynamic‐MRTF/SRF cascade, thereby influencing actin cytoskeleton organization and SRF activity. Additionally, circadian clock exerts precise temporal control over adipogenic differentiation. BMAL1 orchestrates the transcriptional regulation of genes associated with the actin dynamic‐myocardin‐related transcription factor/serum response factor (MRTF/SRF) cascade, thereby influencing the organization of the actin cytoskeleton and modulating SRF activity. This regulatory mechanism holds pivotal importance in the transformation of adipogenic progenitor cells into mature adipocytes, the process of beige adipogenesis, and the enhancement of thermogenic capacity.130 In summary, circadian clocks play a vital and multifaceted role in both the functions and differentiation of adipocytes, underscoring their significance in the realm of adipose tissue physiology and the broader context of systemic energy homeostasis.

Intestinal epithelial cells, a single layer of tightly interconnected columnar cells, form the primary defensive barrier of the intestinal mucosa.131 Circadian clock regulates cellular processes in intestinal epithelial cells including growth, proliferation, differentiation, and cell signaling.132 Of particular note, BMAL1 plays a central role in the regulation of intestinal epithelial cell regeneration by modulating cytokine activity, cell cycle progression, and cell proliferation dynamics. It is worth highlighting that knockdown of Bmal1 results in the disruption of the rhythmic pattern of cellular proliferation.133, 134 Furthermore, TNF cytokine signaling and P21 emerge as key contributors to BMAL1‐mediated cell proliferation control. The former fosters proliferation among epithelial precursors, thereby facilitating the replacement of damaged cells, while the latter assumes a pivotal role in regulating diurnal rhythms in cell production.134 The epithelial clock also exerts regulatory control over essential molecules such as Wee1, cyclins D1, and E, all of which are indispensable for the generation and maintenance of circadian rhythms in the cell cycle progression of intestinal epithelial cells.135 Additionally, intestinal stem cells, vital for the continuous replenishment of differentiated epithelial cells, are influenced by clock components including BMAL1 or CRY. These components modulate transcripts linked to regeneration and stem cell signaling, encompassing pathways like Wnt and Hippo signaling within the intestinal epithelium.136

The circadian clock serves as a key regulator in managing both endogenous substances and xenobiotics within intestinal epithelial cells, highlighting its essential role in cellular processing and homeostasis. Intestinal BMAL1 transcriptionally enhances DGAT2 expression, an enzyme crucial for triacylglycerol synthesis, by binding to an E‐box in its promoter, thereby facilitating dietary fat absorption in the gut.137 Intestinal exporter MRP2, which plays a key role in the disposition and elimination of various drugs, is also impacted by the intestinal circadian clock. Loss of Bmal1 in intestinal epithelial cells diminishes the expression and transport activity of MRP2, as well as their diurnal rhythms, affecting the pharmacokinetics and toxicity of medications like methotrexate.138

NF‐κB signaling pathway, comprising a family of inducible transcription factors, is a pivotal regulator of inflammatory responses.140 Circadian clock modulates various aspects of the NF‐κB signaling pathway targets (Figure 5).141 Clock factors form an intricate network with NF‐κB proteins, influencing their function and interaction. REV‐ERBα, a key clock component, regulates the levels of p65, a subunit of NF‐κB, orchestrating the expressions of inflammatory genes and the release of cytokines and chemokines through three distinct mechanisms. First, Rev‐erbα suppresses the Toll‐Like Receptor 4, thereby blocking upstream pro‐inflammatory signals in the NF‐κB pathway.142 Second, REV‐ERBα disminishes the nuclear level of p65 protein, inhibiting its transcriptional activation of target genes. Third, REV‐ERBα directly binds to the promoters of inflammatory genes (e.g., [NOD‐, LRR‐ and pyrin domain‐containing protein 3]Nlrp3, Il‐1β, and Il‐6), suppressing their transcription and expressions. Multiple mechanisms are proposed for the regulation of NF‐κB signaling by the clock protein BMAL1. BMAL1 regulates NF‐κB/NLRP3 axis via REV‐ERBα, which acts as a repressor of NF‐κB/NLRP3. Supporting this, BMAL1 influences the timing of gene expression in response to inflammatory activation by modulating the epigenetic status of enhancers, partly through the regulation of REV‐ERBs and eRNA transcription in macrophages.143 Biochemical and biophysical analyses have shown that p65 subunit of NF‐κB binds to the transactivation domain of BMAL1 through protein–protein interaction,144 with additional mechanisms including an increase in the phosphorylation of IκB.144 CLOCK directly interacts with p65 protein, and CLOCK overexpression leads to an increase in specific phosphorylated and acetylated transcriptionally active forms of p65.145 CRY1, another clock component, binds to adenylyl cyclase, limiting cAMP production and inactivating protein kinase A, which results in NF‐κB activation through phosphorylation of p65 at S276.146 These studies provide a comprehensive mechanistic framework for understanding the intricate relationship between the circadian clock and NF‐κB signaling.

MAPK signaling pathways are composed of three families including extracellular‐signal‐regulated kinase (ERK), c‐Jun NH2‐terminal kinase (JNK), and p38. Clock factors physically and/or genetically interact with MAPKs to regulate inflammatory responses. BMAL1 decreases ERK phosphorylation in chondrocytes147 and inactivates p38 phosphorylation after traumatic brain injury.78 REV‐ERBα suppresses osteoblast differentiation by inhibiting the p38 MAPK pathway,148 while ROR‐γ‐knockdown downregulates p38 MAPK expression in hepatocytes.149 These regulatory mechanisms are attributed to circadian rhythms in the expression of MAPK genes and/or proteins such as ERK and MAPKs (upstream of p38 and JNK kinases).150

JAK‐STAT signaling pathway is critical for orchestrating the immune system. The circadian clock modulates JAK‐STAT signaling pathway via two separable mechanisms. First, REV‐ERBα abrogates JAK/STAT3 signaling pathway as evidenced by reduced phosphorylated JAK1, JAK2, and STAT3. This effect was attained by up‐regulating the suppressor of cytokine signaling 3, an inhibitor of the JAK/STAT3.151 In addition to the core clock genes, miR‐279 acting as an effector of clock‐controlled behavioral output, targets Upd (the ortholog of the JAK/STAT ligand) to circadian behavioral output.152 Consequently, STAT92E and phosphorylated HOP (JAK/STAT components) are expressed in a circadian manner.153 The regulation of JAK/STAT3 by circadian clock bears significance for the etiology of inflammatory and autoimmune disorders.

Cancer remains a leading cause of mortality and a significant barrier to extending life expectancy. Emerging research increasingly implicates disruptions in circadian rhythms as significant factors in the onset and progression of various cancers. For instance, shift workers exhibit an approximately 10% heightened risk of developing breast cancer.170 Furthermore, mice with chronic circadian disruption are more likely to develop HCC (hepatocellular carcinoma), likely due to metabolic and immunologic changes.171 Additionally, chronic jet lag exacerbates tumor‐induced inflammation in the mouse liver and hypothalamus, which may contribute to cancer‐related cognitive impairments and an unfavorable prognosis.172 In a contrasting perspective, the levels of specific clock genes, namely, PER1, PER2, PER3, and CRY2, are notably diminished in cancerous tissues, compared to their noncancerous counterparts. This reduction is primarily ascribed to promoter methylation and overexpression of EZH2, a gene integral to epigenetic regulation.173

Alterations in the components of the circadian clock, including BMAL1, PER1/2, and CRY1/2, have been correlated with an increased risk of cancer development. Specifically, BMAL1 is critical in hematologic malignancies, where its inactivation has been linked to the advancement of cancers.174Bmal1 knockout mice show a heightened susceptibility to colitis‐associated colorectal cancer.160 Mice with hepatocyte Bmal1 deficiency have a protective effect in high‐fat diet‐ and diethylnitrosamine‐induced hepatocellular carcinoma in mice.175 The repressive arms of the core circadian clock, including PER1 and PER2, are also implicated in cancer. Reduced expression of these genes has been associated with shorter survival in patients with glioma and gastric cancer.176 These findings highlight the intricate connection between circadian rhythms and cancer, emphasizing the need for further research to understand how circadian disruptions contribute to cancer development and progression.

Inflammation plays a pivotal role in the regulatory effects of the circadian clock on cancer pathogenesis.177, 178, 179 The activation, trafficking, and infiltration of immune cells into tumors are largely contingent on the release and presentation of cancer cell antigens, with inflammation being a central mediator in this mechanism. For instance, rhythmic expression of Bmal1 regulates the numbers of antigen‐specific DCs and the volume of tumors following tumor engraftment at different time points (ZT9 or ZT21).180 In a melanoma mouse model, chronic jet lag leads to significant immunosuppression within the tumor microenvironment, accelerating tumor growth. This effect is attributed to the elimination of daily variations in macrophage counts and a reduced ratio of M1 to M2 macrophages in the tumor.73 Additionally, Bmal1 deficiency in mice results in a decrease in Breg+ PDL1+ cell numbers among intestinal intraepithelial lymphocytes and induces the death of activated CD4+ T cells, thereby promoting the progression of colitis‐associated colorectal cancer.160 Chronic shift‐lag conditions disrupt both the circadian expressions of clock genes and the cytolytic activities of NK cells, potentially fostering tumor growth in the lung.156 Furthermore, various regulators, such as cyclin‐dependent kinases, suppressor of mother against decapentaplegic (SMAD) family member 4, chemokines, vascular endothelial growth factor A, and telomerase reverse transcriptase, have been implicated in the influence of clock genes on cancer development.176 Given these insights, it is crucial to explore novel therapeutic approaches targeting cancer cells, leveraging the circadian rhythmic regulation of cancer biology. Such strategies could offer more effective and tailored treatments, potentially improving patient outcomes by aligning therapy with the body's natural circadian rhythms.

All data are available upon request.