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1. Repeated evolution of circadian clock dysregulation in cavefish populations

   https://doi.org/10.1371/journal.pgen.1009642  

   Mack, Katya L.; Jaggard, James B.; Persons, Jenna L.; Roback, Emma Y.; Passow, Courtney N.; Stanhope, Bethany A.; Ferrufino, Estephany; Tsuchiya, Dai; Smith, Sarah E.; Slaughter, Brian D.; et al (July 2021, PLOS Genetics)

   Fay, Justin C. (Ed.)

   Circadian rhythms are nearly ubiquitous throughout nature, suggesting they are critical for survival in diverse environments. Organisms inhabiting largely arrhythmic environments, such as caves, offer a unique opportunity to study the evolution of circadian rhythms in response to changing ecological pressures. Populations of the Mexican tetra, Astyanax mexicanus , have repeatedly invaded caves from surface rivers, where individuals must contend with perpetual darkness, reduced food availability, and limited fluctuations in daily environmental cues. To investigate the molecular basis for evolved changes in circadian rhythms, we investigated rhythmic transcription across multiple independently-evolved cavefish populations. Our findings reveal that evolution in a cave environment has led to the repeated disruption of the endogenous biological clock, and its entrainment by light. The circadian transcriptome shows widespread reductions and losses of rhythmic transcription and changes to the timing of the activation/repression of core-transcriptional clock. In addition to dysregulation of the core clock, we find that rhythmic transcription of the melatonin regulator aanat2 and melatonin rhythms are disrupted in cavefish under darkness. Mutants of aanat2 and core clock gene rorca disrupt diurnal regulation of sleep in A . mexicanus , phenocopying circadian modulation of sleep and activity phenotypes of cave populations. Together, these findings reveal multiple independent mechanisms for loss of circadian rhythms in cavefish populations and provide a platform for studying how evolved changes in the biological clock can contribute to variation in sleep and circadian behavior. 

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2. Keeping time in the dark: Potato diel and circadian rhythmic gene expression reveals tissue‐specific circadian clocks

   https://doi.org/10.1002/pld3.425  

   Hoopes, Genevieve M.; Zarka, Daniel; Feke, Ann; Acheson, Kaitlyn; Hamilton, John P.; Douches, David; Buell, C. Robin; Farré, Eva M. (July 2022, Plant Direct)

   Abstract The circadian clock is an internal molecular oscillator and coordinates numerous physiological processes through regulation of molecular pathways. Tissue‐specific clocks connected by mobile signals have previously been found to run at different speeds inArabidopsis thalianatissues. However, tissue variation in circadian clocks in crop species is unknown. In this study, leaf and tuber global gene expression in cultivated potato under cycling and constant environmental conditions was profiled. In addition, we used a circadian‐regulated luciferase reporter construct to study tuber gene expression rhythms. Diel and circadian expression patterns were present among 17.9% and 5.6% of the expressed genes in the tuber. Over 500 genes displayed differential tissue specific diel phases. Intriguingly, few core circadian clock genes had circadian expression patterns, while all such genes were circadian rhythmic in cultivated tomato leaves. Furthermore, robust diel and circadian transcriptional rhythms were observed among detached tubers. Our results suggest alternative regulatory mechanisms and/or clock composition is present in potato, as well as the presence of tissue‐specific independent circadian clocks. We have provided the first evidence of a functional circadian clock in below‐ground storage organs, holding important implications for other storage root and tuberous crops. 

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3. Investigating the Roles for Essential Genes in the Regulation of the Circadian Clock in Synechococcus elongatus Using CRISPR Interference

   https://doi.org/10.1177/07487304241228333  

   Boodaghian, Nouneh; Park, Hyunsook; Cohen, Susan_E (February 2024, Journal of Biological Rhythms)

   Circadian rhythms are found widely throughout nature where cyanobacteria are the simplest organisms, in which the molecular details of the clock have been elucidated. Circadian rhythmicity in cyanobacteria is carried out via the KaiA, KaiB, and KaiC core oscillator proteins that keep ~24 h time. A series of input and output proteins—CikA, SasA, and RpaA—regulate the clock by sensing environmental changes and timing rhythmic activities, including global rhythms of gene expression. Our previous work identified a novel set of KaiC-interacting proteins, some of which are encoded by genes that are essential for viability. To understand the relationship of these essential genes to the clock, we applied CRISPR interference (CRISPRi) which utilizes a deactivated Cas9 protein and single-guide RNA (sgRNA) to reduce the expression of target genes but not fully abolish their expression to allow for survival. Eight candidate genes were targeted, and strains were analyzed by quantitative real-time PCR (qRT-PCR) for reduction of gene expression, and rhythms of gene expression were monitored to analyze circadian phenotypes. Strains with reduced expression of SynPCC7942_0001, dnaN, which encodes for the β-clamp of the replicative DNA polymerase, or SynPCC7942_1081, which likely encodes for a KtrA homolog involved in K⁺ transport, displayed longer circadian rhythms of gene expression than the wild type. As neither of these proteins have been previously implicated in the circadian clock, these data suggest that diverse cellular processes, DNA replication and K⁺ transport, can influence the circadian clock and represent new avenues to understand clock function. 

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4. Methamphetamine alters the circadian oscillator and its couplings on multiple scales in Per1/2/3 knockout mice

   https://doi.org/10.1093/pnasnexus/pgaf070  

   Barnes, Samuel_J_K; Alanazi, Mansour; Yamazaki, Shin; Stefanovska, Aneta; Aeschbach, ed., Daniel (March 2025, PNAS Nexus)

   Abstract Disruptions to circadian rhythms in mammals are associated with alterations in their physiological and mental states. Circadian rhythms are currently analyzed in the time domain using approaches such as actograms, thus failing to appreciate their time-localized characteristics, time-varying nature and multiscale dynamics. In this study, we apply time-resolved analysis to investigate behavioral rhythms in Per1/2/3 knockout (KO) mice and their changes following methamphetamine administration, focusing on circadian (around 24 h), low-frequency ultradian (around 7 h), high-frequency ultradian (around 30 min), and circabidian (around 48 h) oscillations. In the absence of methamphetamine, Per1/2/3 KO mice in constant darkness exhibited a dominant, ∼7 h oscillation. We demonstrate that methamphetamine exposure restores the circadian rhythm, although the frequency of the methamphetamine sensitive circadian oscillator varied considerably compared to the highly regular wild-type circadian rhythm. Additionally, methamphetamine increased multiscale activity and induced a circabidian oscillation in the Per1/2/3 KO mice. The information transfer between oscillatory modes, with frequencies around circadian, low-frequency ultradian and high-frequency ultradian activity, due to their mutual couplings, was also investigated. For Per1/2/3 KO mice in constant darkness, the most prevalent coupling was between low and high-frequency ultradian activity. Following methamphetamine administration, the coupling between the circadian and high-frequency ultradian activity became dominant. In each case, the direction of information transfer was between the corresponding phases from the slower to faster oscillations. The time-varying nature of the circadian rhythm exhibited in the absence of Per1/2/3 genes and following methamphetamine administration may have profound implications for health and disease. 

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5. The role of temperature on the development of circadian rhythms in honey bee workers

   https://doi.org/10.7717/peerj.17086  

   Giannoni-Guzmán, Manuel A; Perez_Claudio, Eddie; Aleman-Rios, Janpierre; Diaz_Hernandez, Gabriel; Perez_Torres, Melina; Melendez_Moreno, Alexander; Loubriel, Darimar; Moore, Darrell; Giray, Tugrul; Agosto-Rivera, Jose L (January 2024, PeerJ)

   Circadian rhythms in honey bees are involved in various processes that impact colony survival. For example, young nurses take care of the brood constantly throughout the day and lack circadian rhythms. At the same time, foragers use the circadian clock to remember and predict food availability in subsequent days. Previous studies exploring the ontogeny of circadian rhythms of workers showed that the onset of rhythms is faster in the colony environment (~2 days) than if workers were immediately isolated after eclosion (7–9 days). However, which specific environmental factors influenced the early development of worker circadian rhythms remained unknown. We hypothesized that brood nest temperature plays a key role in the development of circadian rhythmicity in young workers. Our results show that young workers kept at brood nest-like temperatures (33–35 °C) in the laboratory develop circadian rhythms faster and in greater proportion than bees kept at lower temperatures (24–26 °C). In addition, we examined if the effect of colony temperature during the first 48 h after emergence is sufficient to increase the rate and proportion of development of circadian rhythmicity. We observed that twice as many individuals exposed to 35 °C during the first 48 h developed circadian rhythms compared to individuals kept at 25 °C, suggesting a critical developmental period where brood nest temperatures are important for the development of the circadian system. Together, our findings show that temperature, which is socially regulated inside the hive, is a key factor that influences the ontogeny of circadian rhythmicity of workers. 

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