Circadian Expression Patterns Research Articles

Nighttime-specific differential gene expression in suprachiasmatic nucleus and habenula is associated with resilience to chronic social stress

The molecular mechanisms that link stress and biological rhythms still remain unclear. The habenula (Hb) is a key brain region involved in regulating diverse types of emotion-related behaviours while the suprachiasmatic nucleus (SCN) is the body’s central clock. To investigate the effects of chronic social stress on transcription patterns, we performed gene expression analysis in the Hb and SCN of stress-naïve and stress-exposed mice. Our analysis revealed a large number of differentially expressed genes and enrichment of synaptic and cell signalling pathways between resilient and stress-naïve mice at zeitgeber 16 (ZT16) in both the Hb and SCN. This transcriptomic signature was nighttime-specific and observed only in stress-resilient mice. In contrast, there were relatively few differences between the stress-susceptible and stress-naïve groups across time points. Our results reinforce the functional link between circadian gene expression patterns and differential responses to stress, thereby highlighting the importance of temporal expression patterns in homoeostatic stress responses.

Circadian cilia transcriptome in mouse brain across physiological and pathological states

Primary cilia are dynamic sensory organelles that continuously undergo structural modifications in response to environmental and cellular signals, many of which exhibit rhythmic patterns. Building on our previous findings of rhythmic cilia-related gene expression in diurnal primates (baboon), this study extends the investigation to the nocturnal mouse brain to identify circadian patterns of cilia gene expression across brain regions. We used computational techniques and transcriptomic data from four publicly available databases, to examine the circadian expression of cilia-associated genes within six brain areas: brainstem, cerebellum, hippocampus, hypothalamus, striatum, and suprachiasmatic nucleus. Our analysis reveals that a substantial proportion of cilia transcripts exhibit circadian rhythmicity across the examined regions, with notable overrepresentation in the striatum, hippocampus, and cerebellum. We also demonstrate region-specific variations in the abundance and timing of circadian cilia genes’ peaks, indicating an adaptation to the distinct physiological roles of each brain region. Additionally, we show that the rhythmic patterns of cilia transcripts are shifted under various physiological and pathological conditions, including modulation of the dopamine system, high-fat diet, and epileptic conditions, indicating the adaptable nature of cilia transcripts’ oscillation. While limited to a few mouse brain regions, our study provides initial insights into the distinct circadian patterns of cilia transcripts and highlights the need for future research to expand the mapping across wider brain areas to fully understand the role of cilia’s spatiotemporal dynamics in brain functions.

Transcriptome Analysis Revealed ZmPTOX1 Is Required for Seedling Development and Stress Tolerance in Maize.

Plant seedling morphogenesis is considerably related to photosynthesis, pigment synthesis, and circadian periodicity during seedling development. We identified and cloned a maize zebra or crossbanding leaves mutant wk3735, which produces pale white kernels and was identified and plays a role in the equilibrium of the Redox state the in/out of ETC by active oxygen scavenging. Interestingly, it produces the zebra leaves during the production of the first seven leaves, which is apparently different from the mutation of homologs AtPTOX in Arabidopsis. It is intriguing to investigate how and why yellow crossbands (zebra leaf phenotype) emerge on leaves. As expected, chlorophyll concentration and photosynthetic efficiency both significantly declined in the yellow sector of wk3735 leaves. Meanwhile, we observed the circadian expression pattern of ZmPTOX1, which was further validated by protein interaction assays of the circadian clock protein TIM1 and ZmPTOX1. The transcriptome data of yellow (muW) and green (muG) sectors of knock-out lines and normal leaves of overexpression lines (OE) at the 5th-leaf seedling stage were analyzed. Zebra leaf etiolated sections exhibit a marked defect in the expression of genes involved in the circadian rhythm and rhythmic stress (light and cold stress) responses than green sections. According to the analysis of co-DEGs of muW vs. OE and muG vs. OE, terms linked to cell repair function were upregulated while those linked to environmental adaptability and stress response were downregulated due to the mutation of ZmPTOX1. Further gene expression level analyses of reactive oxygen species (ROS) scavenging enzymes and detection of ROS deposition indicated that ZmPTOX1 played an essential role in plant stress resistance and ROS homeostasis. The pleiotropic roles of ZmPTOX1 in plant ROS homeostasis maintenance, stress response, and circadian rhythm character may collectively explain the phenotype of zebra leaves during wk3735 seedling development.

Circadian clock shapes diurnal gene expression patterns linked to glucose metabolic processes in Chinese cabbage

The plant circadian clock temporally drives gene expression through the day and coordinates various physiological process with diurnal environmental changes. It is essential to confer plant fitness and competitive advantage to survive and thrive under natural condition by circadian control of gene transcription. Chinese cabbage (Brassica rapa ssp. pekinensis) is an economically important vegetable crop worldwide; however, there is little information concerning its circadian clock system. Here we uncovered that gene expression patterns were affected by circadian oscillators at both transcriptional and post-transcriptional levels in Chinese cabbage. Time-course RNA-seq analyses were conducted on two short-period lines (SPcc-1 and SPcc-2) and two long-period lines (LPcc-1 and LPcc-2) under constant light. We showed that 32.7-50.5% of the genes were regulated by the circadian oscillator and the expression peak of cycling genes appeared even earlier in short-period lines compared to long-period lines. In addition, approximately 250 splicing events showed circadian regulation, of which intron retention (IR) accounted for a large proportion. Rhythmically spliced genes included the clock genes LATE ELONGATED HYOCOTYL (BrLHY), REVEILLE 2 (BrRVE2) and EARLY FLOWERING 3 (BrELF3). We also found that the circadian oscillator could notably influence the diurnal expression patterns of genes that are associated with glucose metabolism via photosynthesis, Calvin cycle and tricarboxylic acid (TCA) cycle at both transcriptional and post-transcriptional levels. Taken together, our results demonstrate that circadian-regulated physiological processes contribute to Chinese cabbage growth and development.

Quantification of circadian rhythms in mammalian lung tissue snapshot data

Healthy mammalian cells have a circadian clock, a gene regulatory network that allows them to schedule their physiological processes to optimal times of the day. When healthy cells turn into cancer cells, the circadian clock often becomes cancer specifically disturbed, so there is an interest in the extraction of circadian features from gene expression data of cancer. This is challenging, as clinical gene expression samples of cancer are snapshot-like and the circadian clock is best examined using gene expression time series. In this study, we obtained lists of intersecting circadian genes in public gene expression time series data of lung tissue of mouse and baboon. We base our circadian gene lists on correlations of gene expression levels of circadian genes, which are closely associated to the phase differences between them. Combining circadian gene expression patterns of diurnal and nocturnal species of different ages provides circadian genes that are also important in healthy and cancerous human lung tissue. We tested the quality of the representation of the circadian clock in our gene lists by PCA-based reconstructions of the circadian times of the mouse and baboon samples. Then we assigned potential circadian times to the human lung tissue samples and find an intact circadian clock in the healthy human lung tissue, but an altered, weak clock in the adjacent cancerous lung tissue.

Dysregulation of circadian clock gene expression patterns in a treatment-resistant animal model of depression.

Circadian rhythm (CR) disturbances are among the most commonly observed symptoms during major depressive disorder, mostly in the form of disrupted sleeping patterns. However, several other measurable parameters, such as plasma hormone rhythms and differential expression of circadian clock genes (ccgs), are also present, often referred to as circadian phase markers. In the recent years, CR disturbances have been recognized as an essential aspect of depression; however, most of the known animal models of depression have yet to be evaluated for their eligibility to model CR disturbances. In this study, we investigate the potential of adrenocorticotropic hormone (ACTH)-treated animals as a disease model for research in CR disturbances in treatment-resistant depression. For this purpose, we evaluate the changes in several circadian phase markers, including plasma concentrations of corticosterone, ACTH, and melatonin, as well as gene expression patterns of 13 selected ccgs at 3 different time points, in both peripheral and central tissues. We observed no impact on plasma corticosterone and melatonin concentrations in the ACTH rats compared to vehicle. However, the expression pattern of several ccgs was affected in the ACTH rats compared to vehicle. In the hippocampus, 10 ccgs were affected by ACTH treatment, whereas in the adrenal glands, 5 ccgs were affected and in the prefrontal cortex, hypothalamus and liver 4 ccgs were regulated. In the blood, only 1 gene was affected. Individual tissues showed changes in different ccgs, but the expression of Bmal1, Per1, and Per2 were most generally affected. Collectively, the results presented here indicate that the ACTH animal model displays dysregulation of a number of phase markers suggesting the model may be appropriate for future studies into CR disturbances.

Fibroblasts as an in vitro model of circadian genetic and genomic studies

Bipolar disorder (BD) is a heritable disorder characterized by shifts in mood that manifest in manic or depressive episodes. Clinical studies have identified abnormalities of the circadian system in BD patients as a hallmark of underlying pathophysiology. Fibroblasts are a well-established in vitro model for measuring circadian patterns. We set out to examine the underlying genetic architecture of circadian rhythm in fibroblasts, with the goal to assess its contribution to the polygenic nature of BD disease risk. We collected, from primary cell lines of 6 healthy individuals, temporal genomic features over a 48 h period from transcriptomic data (RNA-seq) and open chromatin data (ATAC-seq). The RNA-seq data showed that only a limited number of genes, primarily the known core clock genes such as ARNTL, CRY1, PER3, NR1D2 and TEF display circadian patterns of expression consistently across cell cultures. The ATAC-seq data identified that distinct transcription factor families, like those with the basic helix-loop-helix motif, were associated with regions that were increasing in accessibility over time. Whereas known glucocorticoid receptor target motifs were identified in those regions that were decreasing in accessibility. Further evaluation of these regions using stratified linkage disequilibrium score regression analysis failed to identify a significant presence of them in the known genetic architecture of BD, and other psychiatric disorders or neurobehavioral traits in which the circadian rhythm is affected. In this study, we characterize the biological pathways that are activated in this in vitro circadian model, evaluating the relevance of these processes in the context of the genetic architecture of BD and other disorders, highlighting its limitations and future applications for circadian genomic studies.

Lack of Bmal1 leads to changes in rhythmicity and impairs motivation towards natural stimuli.

Maintaining proper circadian rhythms is essential for coordinating biological functions in mammals. This study investigates the effects of daily arrhythmicity using Bmal1-knockout (KO) mice as a model, aiming to understand behavioural and motivational implications. By employing a new mathematical analysis based on entropy divergence, we identified disrupted intricate activity patterns in mice derived by the complete absence of BMAL1 and quantified the difference regarding the activity oscillation's complexity. Changes in locomotor activity coincided with disturbances in circadian gene expression patterns. Additionally, we found a dysregulated gene expression profile particularly in brain nuclei like the ventral striatum, impacting genes related to reward and motivation. Further investigation revealed that arrhythmic mice exhibited heightened motivation for food and water rewards, indicating a link between circadian disruptions and the reward system. This research sheds light on how circadian clock alterations impact the gene expression regulating the reward system and how this, in turn, can lead to altered seeking behaviour and motivation for natural rewards. In summary, the present study contributes to our understanding of how reward processing is under the regulation of circadian clock machinery.

Severe asthmatic airways have distinct circadian clock gene expression pattern associated with WNT signaling

Severe asthmatic airways have distinct circadian clock gene expression pattern associated with WNT signaling

A glial circadian gene expression atlas reveals cell type and disease-specific reprogramming in response to amyloid pathology or aging.

While circadian rhythm disruption may promote neurodegenerative disease, how aging and neurodegenerative pathology impact circadian gene expression patterns in different brain cell types is unknown. Here, we used translating ribosome affinity purification methods to define the circadian translatomes of astrocytes, microglia, and bulk cerebral cortex, in healthy mouse brain and in the settings of amyloid-beta plaque pathology or aging. Our data reveal that glial circadian translatomes are highly cell type-specific and exhibit profound, context-dependent reprogramming of rhythmic transcripts in response to amyloid pathology or aging. Transcripts involved in glial activation, immunometabolism, and proteostasis, as well as nearly half of all Alzheimer Disease (AD)-associated risk genes, displayed circadian oscillations, many of which were altered by pathology. Amyloid-related differential gene expression was also dependent on time of day. Thus, circadian rhythms in gene expression are cell- and context dependent and provide important insights into glial gene regulation in health, AD, and aging.

Clock-dependent chromatin accessibility rhythms regulate circadian transcription.

Chromatin organization plays a crucial role in gene regulation by controlling the accessibility of DNA to transcription machinery. While significant progress has been made in understanding the regulatory role of clock proteins in circadian rhythms, how chromatin organization affects circadian rhythms remains poorly understood. Here, we employed ATAC-seq (Assay for Transposase-Accessible Chromatin with Sequencing) on FAC-sorted Drosophila clock neurons to assess genome-wide chromatin accessibility at dawn and dusk over the circadian cycle. We observed significant oscillations in chromatin accessibility at promoter and enhancer regions of hundreds of genes, with enhanced accessibility either at dusk or dawn, which correlated with their peak transcriptional activity. Notably, genes with enhanced accessibility at dusk were enriched with E-box motifs, while those more accessible at dawn were enriched with VRI/PDP1-box motifs, indicating that they are regulated by the core circadian feedback loops, PER/CLK and VRI/PDP1, respectively. Further, we observed a complete loss of chromatin accessibility rhythms in per01 null mutants, with chromatin consistently accessible at both dawn and dusk, underscoring the critical role of Period protein in driving chromatin compaction during the repression phase at dawn. Together, this study demonstrates the significant role of chromatin organization in circadian regulation, revealing how the interplay between clock proteins and chromatin structure orchestrates the precise timing of biological processes throughout the day. This work further implies that variations in chromatin accessibility might play a central role in the generation of diverse circadian gene expression patterns in clock neurons.

Mice lacking DIO3 exhibit sex-specific alterations in circadian patterns of corticosterone and gene expression in metabolic tissues

Disruption of circadian rhythms is associated with neurological, endocrine and metabolic pathologies. We have recently shown that mice lacking functional type 3 deiodinase (DIO3), the enzyme that clears thyroid hormones, exhibit a phase shift in locomotor activity, suggesting altered circadian rhythm. To better understand the physiological and molecular basis of this phenotype, we used Dio3+/+ and Dio3-/- mice of both sexes at different zeitgeber times (ZTs) and analyzed corticosterone and thyroxine (T4) levels, hypothalamic, hepatic, and adipose tissue expression of clock genes, as well as genes involved in the thyroid hormone action or physiology of liver and adipose tissues. Wild type mice exhibited sexually dimorphic circadian patterns of genes controlling thyroid hormone action, including Dio3. Dio3-/- mice exhibited altered hypothalamic expression of several clock genes at ZT12, but did not disrupt the overall circadian profile. Expression of clock genes in peripheral tissues was not disrupted by Dio3 deficiency. However, Dio3 loss in liver and adipose tissues disrupted circadian profiles of genes that determine tissue thyroid hormone action and physiology. We also observed circadian-specific changes in serum T4 and corticosterone as a result of DIO3 deficiency. The circadian alterations manifested sexual dimorphism. Most notable, the time curve of serum corticosterone was flattened in Dio3-/- females. We conclude that Dio3 exhibits circadian variations, influencing the circadian rhythmicity of thyroid hormone action and physiology in liver and adipose tissues in a sex-specific manner. Circadian disruptions in tissue physiology may then contribute to the metabolic phenotypes of DIO3-deficient mice.

Deficiency of circadian clock gene Bmal1 exacerbates noncanonical inflammasome-mediated pyroptosis and lethality via Rev-erbα-C/EBPβ-SAA1 axis

Circadian arrhythmia has been linked to increased susceptibility to multiple inflammatory diseases, such as sepsis. However, it remains unclear how disruption of the circadian clock modulates molecular aspects of innate immune responses, including inflammasome signaling. Here, we examined the potential role of the circadian clock in inflammasome-mediated responses through myeloid-specific deletion of BMAL1, a master circadian clock regulator. Intriguingly, Bmal1 deficiency significantly enhanced pyroptosis of macrophages and lethality of mice under noncanonical inflammasome-activating conditions but did not alter canonical inflammasome responses. Transcriptome analysis of enriched peritoneal myeloid cells revealed that Bmal1 deficiency led to a marked reduction in Rev-erbα expression at steady state and a significant increase in serum amyloid A1 (SAA1) expression upon poly(I:C) stimulation. Notably, we found that the circadian regulator Rev-erbα is critical for poly(I:C)- or interferon (IFN)-β-induced SAA1 production, resulting in the circadian oscillation pattern of SAA1 expression in myeloid cells. Furthermore, exogenously applied SAA1 markedly increased noncanonical inflammasome-mediated pyroptosis of macrophages and lethality of mice. Intriguingly, our results revealed that type 1 IFN receptor signaling is needed for poly(I:C)- or IFN-β-induced SAA1 production. Downstream of the type 1 IFN receptor, Rev-erbα inhibited the IFN-β-induced association of C/EBPβ with the promoter region of Saa1, leading to the reduced transcription of Saa1 in macrophages. Bmal1-deficient macrophages exhibited enhanced binding of C/EBPβ to Saa1. Consistently, the blockade of Rev-erbα by SR8278 significantly increased poly(I:C)-stimulated SAA1 transcription and noncanonical inflammasome-mediated lethality in mice. Collectively, our data demonstrate a potent suppressive effect of the circadian clock BMAL1 on the noncanonical inflammasome response via the Rev-erbα-C/EBPβ-SAA1 axis.

Comprehensive analysis of circadian protein expression patterns in healthy adults

Comprehensive analysis of circadian protein expression patterns in healthy adults

Circadian protein expression patterns in healthy young adults

ObjectivesTo explore how the blood plasma proteome fluctuates across the 24-hour day and identify a subset of proteins that show endogenous circadian rhythmicity. MethodsPlasma samples from 17 healthy adults were collected hourly under controlled conditions designed to unmask endogenous circadian rhythmicity; in a subset of 8 participants, we also collected samples across a day on a typical sleep-wake schedule. A total of 6916 proteins were analyzed from each sample using the SomaScan aptamer-based multiplexed platform. We used differential rhythmicity analysis based on a cosinor model with mixed effects to identify a subset of proteins that showed circadian rhythmicity in their abundance. ResultsOne thousand and sixty-three (15%) proteins exhibited significant daily rhythmicity. Of those, 431 (6.2%) proteins displayed consistent endogenous circadian rhythms on both a sleep-wake schedule and under controlled conditions: it included both known and novel proteins. When models were fitted with two harmonics, an additional 259 (3.7%) proteins exhibited significant endogenous circadian rhythmicity, indicating that some rhythmic proteins cannot be solely captured by a simple sinusoidal model. Overall, we found that the largest number of proteins had their peak levels in the late afternoon/evening, with another smaller group peaking in the early morning. ConclusionsThis study reveals that hundreds of plasma proteins exhibit endogenous circadian rhythmicity in humans. Future analyses will likely reveal novel physiological pathways regulated by circadian clocks and pave the way for improved diagnosis and treatment for patients with circadian disorders and other pathologies. It will also advance efforts to include knowledge about time-of-day, thereby incorporating circadian medicine into personalized medicine.

Novel Insights into the Circadian Rhythms Based on Long Noncoding and Circular RNA Profiling.

Circadian rhythm disorders pose major risks to human health and animal production activity, and the hypothalamus is the center of circadian rhythm regulation. However, the epigenetic regulation of circadian rhythm based on farm animal models has been poorly investigated. We collected chicken hypothalamus samples at seven time points in one light/dark cycle and performed long noncoding RNA (lncRNA), circular RNA (circRNA), and mRNA sequencing to detect biomarkers associated with circadian rhythm. We enhanced the comprehensive expression profiling of ncRNAs and mRNAs in the hypothalamus and found two gene sets (circadian rhythm and retinal metabolism) associated with the light/dark cycle. Noncoding RNA networks with circadian expression patterns were identified by differential expression and circadian analysis was provided that included 38 lncRNAs, 15 circRNAs, and 200 candidate genes. Three lncRNAs (ENSGALT00000098661, ENSGALT00000100816, and MSTRG.16980.1) and one circRNA (novel_circ_010168) in the ncRNA-mRNA regulatory network were identified as key molecules influencing circadian rhythm by regulating AOX1 in retinal metabolism. These ncRNAs were predicted to be related to pernicious anemia, gonadal, eye disease and other disorders in humans. Together, the findings of this study provide insights into the epigenetic mechanisms of circadian rhythm and reveal AOX1 as a promising target of circadian rhythm regulation.

Period circadian regulator 2-mediated steroid hormone synthesis by regulating transcription of steroidogenic acute regulatory protein in porcine granulosa cells.

Steroidogenesis is associated with circadian clock genes. However, the regulation of steroid hormone production in sow granulosal cells by Per2, a crucial circadian regulator, remains unexplored. In this study, we have identified the presence of Per2 in ovarian granulosa cells and have observed its circadian expression pattern. Employing siRNA to interfere with Per2 expression, our investigation revealed that Per2 knockdown notably elevated progesterone (P4) levels along with increasing the expression of StAR but interference of Per2 did not alter the rhythm of clock-related gene (Bmal1, Clock, Per1, and Cry1) in granulosa cells. Subsequent mechanistic analysis showed that Per2 formed complexes with PPARγ and interference with Per2 promoted the formation of the PPARγ:RXRα heterodimer. Importantly, we uncovered that PPARγ:RXRα heterodimer could control the expression of StAR via direct peroxisome proliferator response element binding to its promoter to regulate its activity, and knockdown of Per2 promoted the transcription of StAR via increasing the binding of PPARγ:RXRα ligands. Altogether, these findings indicated a noncanonical role of Per2 in controlling PPARγ:RXRα binding to regulate transcription of StAR and progesterone synthesis, thus revealing potential avenues of pharmacological and therapeutic intervention.

Molecular characterization and dietary regulation of the forkhead box O3 (foxo3a and foxo3b) in grass carp (Ctenopharyngodon idella)

The forkhead box O3 (foxo3) regulates the target genes expression involved in regulating cell cycle and biological metabolism. In the study, foxo3a and foxo3b of grass carp (Ctenopharyngodon idella) were cloned. The foxo3a cDNA have a complete open reading frame (ORF) spanning 1431 bp, which encodes a polypeptide of 476 amino acids. The foxo3b cDNA contains a complete ORF encompassing 1959 bp, encoding a protein consisting of 652 amino acid residues. The analysis of genetic codon preference and phylogenetic relationships revealed that the foxo3a and foxo3b genes in grass carp are closely related to the counterparts of Danio rerio and Megalobrama amblycephala. Tissue expression analysis showed that foxo3a expression reached the maximum in the brain while foxo3b expression in the muscle being the highest. Circadian expression pattern showed that, foxo3a expression presented a periodic upward trend from 3:00–9:00, 12:00–18:00 and 21:00–24:00, and the maximum level of foxo3b was detected at 9:00. Moreover, dietary regulation of foxo3a and foxo3b expression were performed in our research. In the feeding trial with different protein sources, the groups of fishmeal, rapeseed meal and soybean meal are in contrast to each other. Foxo3a expression was significantly higher in fish fed with rapeseed meal compared to those fed with soybean meal and fishmeal at 14th and 21st days of the experiment. Foxo3b expression was highest in the fishmeal group, followed by rapeseed meal, while it was lowest in soybean meal at 14th and 28th days. Analysis of the effect of dietary alanine-glutamine dipeptide on foxo3 expression in grass carp revealed that the expression of foxo3a gradually increased with increasing dietary Ala-Gln dipeptide concentration and reached its highest level at a concentration of 1.5%. Conversely, foxo3b expression significantly decreased when the dipeptide concentration reached 2%. The findings of this study may provide new insights into the molecular properties, spatiotemporal dynamics, and nutritional regulation of foxo3 in fish.

The WD40 domain-containing protein Ehd5 positively regulates flowering in rice (Oryza sativa).

Heading date (flowering time), which greatly influences regional and seasonal adaptability in rice (Oryza sativa), is regulated by many genes in different photoperiod pathways. Here, we characterized a heading date gene, Early heading date 5 (Ehd5), using a modified bulked segregant analysis method. The ehd5 mutant showed late flowering under both short-day and long-day conditions, as well as reduced yield, compared to the wild type. Ehd5, which encodes a WD40 domain-containing protein, is induced by light and follows a circadian rhythm expression pattern. Transcriptome analysis revealed that Ehd5 acts upstream of the flowering genes Early heading date 1 (Ehd1), RICE FLOWERING LOCUS T 1 (RFT1), and Heading date 3a (Hd3a). Functional analysis showed that Ehd5 directly interacts with Rice outermost cell-specific gene 4 (Roc4) and Grain number, plant height, and heading date 8 (Ghd8), which might affect the formation of Ghd7-Ghd8 complexes, resulting in increased expression of Ehd1, Hd3a and RFT1. Taken together, these results demonstrate that Ehd5 functions as a positive regulator of rice flowering and provide insight into the molecular mechanisms underlying heading date.

Soybean Calmodulin-Binding Transcription Activators, GmCAMTA2 and GmCAMTA8, Coordinate the Circadian Regulation of Developmental Processes and Drought Stress Responses.

The calmodulin-binding transcription activators (CAMTAs) mediate transcriptional regulation of development, growth, and responses to various environmental stresses in plants. To understand the biological roles of soybean CAMTA (GmCAMTA) family members in response to abiotic stresses, we characterized expression patterns of 15 GmCAMTA genes in response to various abiotic stresses. The GmCAMTA genes exhibited distinct circadian regulation expression patterns and were differently expressed in response to salt, drought, and cold stresses. Interestingly, the expression levels of GmCAMTA2, GmCAMTA8, and GmCAMTA12 were higher in stem tissue than in other soybean tissues. To determine the roles of GmCAMTAs in the regulation of developmental processes and stress responses, we isolated GmCAMTA2 and GmCAMTA8 cDNAs from soybean and generated Arabidopsis overexpressing transgenic plants. The GmCAMTA2-OX and GmCAMTA8-OX plants showed hypersensitivity to drought stress. The water in the leaves of GmCAMTA2-OX and GmCAMTA8-OX plants was lost faster than that in wild-type (WT) plants under drought-stress conditions. In addition, stress-responsive genes were down-regulated in the GmCAMTA2-OX and GmCAMTA8-OX plants under drought stress conditions compared to WT plants. Our results suggest that GmCAMTA2 and GmCAMTA8 genes are regulated by circadian rhythms and function as negative regulators in development and drought stress responses.