Fluctuating methylation clocks for cell lineage tracing at high temporal resolution in human tissues

Molecular clocks that record cell ancestry mutate too slowly to measure the short-timescale dynamics of cell renewal in adult tissues. Here, we show that fluctuating DNA methylation marks can be used as clocks in cells where ongoing methylation and demethylation cause repeated ‘flip–flops’ between methylated and unmethylated states. We identify endogenous fluctuating CpG (fCpG) sites using standard methylation arrays and develop a mathematical model to quantitatively measure human adult stem cell dynamics from these data. Small intestinal crypts were inferred to contain slightly more stem cells than the colon, with slower stem cell replacement in the small intestine. Germline APC mutation increased the number of replacements per crypt. In blood, we measured rapid expansion of acute leukemia and slower growth of chronic disease. Thus, the patterns of human somatic cell birth and death are measurable with fluctuating methylation clocks (FMCs).

Circadian Rhythms in the Neuronal Network Timing the Luteinizing Hormone Surge

For 3.5 billion years before electric light was invented in 1879, life on Earth evolved under the pattern of light during the day and darkness during the night. Through evolution, nearly all organisms internalized the temporal rhythm of Earth’s 24-hour rotation and evolved self-sustaining biological clocks with a ~24-hour rhythm. These internal rhythms are called circadian rhythms, and the molecular constituents that generate them are called molecular circadian clocks. Alignment of molecular clocks with the environmental light-dark rhythms optimizes physiology and behavior. This is particularly true for reproductive function, in which seasonal breeders use day-length information to time yearly changes in fertility. However, it is becoming increasingly clear that light-induced disruption of circadian rhythms can negatively impact fertility in non-seasonal breeders as well. In particular, the luteinizing hormone surge promoting ovulation, is sensitive to circadian disruption. In this review, we will summarize our current understanding of the neuronal networks that underlie circadian rhythms and the luteinizing hormone surge.

Melatonin, Clock Genes, and Mammalian Reproduction: What Is the Link?

Physiological processes and behaviors in many mammals are rhythmic. Recently there has been increasing interest in the role of circadian rhythmicity in the control of reproductive function. The circadian rhythm of the pineal hormone melatonin plays a role in synchronizing the reproductive responses of animals to environmental light conditions. There is some evidence that melatonin may have a role in the biological regulation of circadian rhythms and reproduction in humans. Moreover, circadian rhythms and clock genes appear to be involved in optimal reproductive performance. These rhythms are controlled by an endogenous molecular clock within the suprachiasmatic nucleus (SCN) in the hypothalamus, which is entrained by the light/dark cycle. The SCN synchronizes multiple subsidiary oscillators (clock genes) existing in various tissues throughout the body. The basis for maintaining the circadian rhythm is a molecular clock consisting of transcriptional/translational feedback loops. Circadian rhythms and clock genes appear to be involved in optimal reproductive performance. This mini review summarizes the current knowledge regarding the interrelationships between melatonin and the endogenous molecular clocks and their involvement in reproductive physiology (e.g., ovulation) and pathophysiology (e.g., polycystic ovarian syndrome).

Presidential Symposium: Physiological vs. Molecular Clocks, Studies From Mice to Humans

Aging is associated with a functional decline in metabolic, physiological, proliferative, and tissue homeostasis leading to deterioration at the organismal level, and an increased risk for disease and death. Genetic, pharmacological and nutritional interventions have been successfully used to preserve metabolic health, which leads to preserved healthspan and extended longevity. However, the rate at which animals in a population become impaired by age-related frailty and disease is highly variable and several aging clocks that measure different age-modulated processes in the organism are being use as potential markers of the rate of aging. These molecular clocks allow to a more accurate quantification of the biological age of animals. Nevertheless, there is still room for further discussion in terms of the strengths and weaknesses of these biomarkers, in order to probe their biological significance, cellular mechanisms, and epidemiological potential to further explore their long-term benefit of increasing healthspan. This symposium will discuss new approaches to delineate physiological versus molecular clocks based on studies in mice and humans. We will also discuss species-specific metabolic mechanisms based on longitudinal studies in mice, monkeys and humans.

Patterning with clocks and genetic cascades: Segmentation and regionalization of vertebrate versus insect body plans

Oscillatory and sequential processes have been implicated in the spatial patterning of many embryonic tissues. For example, molecular clocks delimit segmental boundaries in vertebrates and insects and mediate lateral root formation in plants, whereas sequential gene activities are involved in the specification of regional identities of insect neuroblasts, vertebrate neural tube, vertebrate limb, and insect and vertebrate body axes. These processes take place in various tissues and organisms, and, hence, raise the question of what common themes and strategies they share. In this article, we review 2 processes that rely on the spatial regulation of periodic and sequential gene activities: segmentation and regionalization of the anterior–posterior (AP) axis of animal body plans. We study these processes in species that belong to 2 different phyla: vertebrates and insects. By contrasting 2 different processes (segmentation and regionalization) in species that belong to 2 distantly related phyla (arthropods and vertebrates), we elucidate the deep logic of patterning by oscillatory and sequential gene activities. Furthermore, in some of these organisms (e.g., the fruit fly Drosophila), a mode of AP patterning has evolved that seems not to overtly rely on oscillations or sequential gene activities, providing an opportunity to study the evolution of pattern formation mechanisms.

A tardigrade in Dominican amber

Tardigrades are a diverse group of charismatic microscopic invertebrates that are best known for their ability to survive extreme conditions. Despite their long evolutionary history and global distribution in both aquatic and terrestrial environments, the tardigrade fossil record is exceedingly sparse. Molecular clocks estimate that tardigrades diverged from other panarthropod lineages before the Cambrian, but only two definitive crown-group representatives have been described to date, both from Cretaceous fossil deposits in North America. Here, we report a third fossil tardigrade from Miocene age Dominican amber. Paradoryphoribius chronocaribbeus gen. et sp. nov. is the first unambiguous fossil representative of the diverse superfamily Isohypsibioidea, as well as the first tardigrade fossil described from the Cenozoic. We propose that the patchy tardigrade fossil record can be explained by the preferential preservation of these microinvertebrates as amber inclusions, coupled with the scarcity of fossiliferous amber deposits before the Cretaceous.

Central and Peripheral Clock Control of Circadian Feeding Rhythms

Many behaviors exhibit ~24-h oscillations under control of an endogenous circadian timing system that tracks time of day via a molecular circadian clock. In the fruit fly, Drosophila melanogaster, most circadian research has focused on the generation of locomotor activity rhythms, but a fundamental question is how the circadian clock orchestrates multiple distinct behavioral outputs. Here, we have investigated the cells and circuits mediating circadian control of feeding behavior. Using an array of genetic tools, we show that, as is the case for locomotor activity rhythms, the presence of feeding rhythms requires molecular clock function in the ventrolateral clock neurons of the central brain. We further demonstrate that the speed of molecular clock oscillations in these neurons dictates the free-running period length of feeding rhythms. In contrast to the effects observed with central clock cell manipulations, we show that genetic abrogation of the molecular clock in the fat body, a peripheral metabolic tissue, is without effect on feeding behavior. Interestingly, we find that molecular clocks in the brain and fat body of control flies gradually grow out of phase with one another under free-running conditions, likely due to a long endogenous period of the fat body clock. Under these conditions, the period of feeding rhythms tracks with molecular oscillations in central brain clock cells, consistent with a primary role of the brain clock in dictating the timing of feeding behavior. Finally, despite a lack of effect of fat body selective manipulations, we find that flies with simultaneous disruption of molecular clocks in multiple peripheral tissues (but with intact central clocks) exhibit decreased feeding rhythm strength and reduced overall food intake. We conclude that both central and peripheral clocks contribute to the regulation of feeding rhythms, with a particularly dominant, pacemaker role for specific populations of central brain clock cells.

Rewinding the molecular clock in the genus Carabus (Coleoptera: Carabidae) in light of fossil evidence and the Gondwana split: A reanalysis

Molecular clocks have become powerful tools given increasing sequencing and fossil resources. However, calibration analyses outcomes depend on the choice of priors. Here, we revisited the seminal dating study published by Andújar and coworkers of the genus Carabus proposing that prior choices need re-evaluation. We hypothesized that reflecting fossil evidence and the Gondwanan split properly significantly rewinds the molecular clock. We re-used the dataset including five mitochondrial and four nuclear DNA fragments with a total length of 7888 nt. Fossil evidence for Oligocene occurrence of Calosoma was considered. Root age was set based on the fossil evidence of Harpalinae ground beetles in the Upper Cretaceous. Paleogene divergence of the outgroup taxa Ceroglossini and Pamborini is introduced as a new prior based on current paleontological and geological literature. The ultrametric time-calibrated tree of the extended nd5 dataset resulted in a median TMRCA Carabus of 53.92 Ma (HPD 95% 45.01–63.18 Ma), roughly 30 Ma older than in the Andújar study. The splits among C. rugosus and C. morbillosus (A), C. riffensis from the European Mesocarabus (B), and Eurycarabus and Nesaeocarabus (C) were dated to 17.58 (12.87–22.85), 24.14 (18.02–30.58), and 21.6 (16.44–27.43) Ma. They were decidedly older than those previously reported (7.48, 10.93, and 9.51 Ma). These changes were driven almost entirely by constraining the Carabidae time-tree root with a Harpalinae amber fossil at ~99 Ma. Utilizing the nd5 dating results of three well-supported Carabus clades as secondary calibration points for the complete MIT-NUC dataset led to a TMRCA of Carabus of 44.72 (37.54–52.22) Ma, compared with 25.16 Ma (18.41–33.04 Ma) in the previous study. Considering fossil evidence for Oligocene Calosoma and Late Cretaceous Harpalini together with the Gondwanan split as a new prior, our new approach supports the origin of genus Carabus in the Eocene. Our results are preliminary because of the heavy reliance on the nd5 gene, and thus will have to be tested with a sufficient set of nuclear markers. Additionally, uncertainties due to dating root age of the tree based on a single fossil and outgroup taxon affect the results. Improvement of the fossil database, particularly in the supertribe Carabitae, is needed to reduce these uncertainties in dating Carabus phylogeny.

Gene networks under circadian control exhibit diurnal organization in primate organs

Mammalian organs are individually controlled by autonomous circadian clocks. At the molecular level, this process is defined by the cyclical co-expression of both core transcription factors and off-target genes across time. While interactions between these molecular clocks are likely necessary for proper homeostasis, these features remain undefined. Here, we utilize integrative analysis of a baboon diurnal transcriptome atlas to characterize the properties of gene networks under circadian control. We found that 53.4% (8,120) of baboon genes are rhythmically expressed body-wide. In addition, >30% of gene-gene interactions exhibit periodic co-expression patterns, with core circadian genes more cyclically co-expressed than others. Moreover, two basic network modes were observed at the systems level: daytime and nighttime mode. Daytime networks were enriched for genes involved in metabolism, while nighttime networks were enriched for genes associated with growth and cellular signaling. A substantial number of diseases only form significant disease modules at either daytime or nighttime. In addition, we found that 216 of 313 genes encoding products that interact with SARS-CoV-2 are rhythmically expressed throughout the body. Importantly, more than 80% of SARS-CoV-2 related genes enriched modules are rhythmically expressed, and have significant network proximities with circadian regulators. Our data suggest that synchronization amongst circadian gene networks is necessary for proper homeostatic functions and circadian regulators have close interactions with SARS-CoV-2 infection.