Molecular clock | EPFL Graph Search

The molecular clock is a figurative term for a technique that uses the mutation rate of biomolecules to deduce the time in prehistory when two or more life forms diverged. The biomolecular data used for such calculations are usually nucleotide sequences for DNA, RNA, or amino acid sequences for proteins. The benchmarks for determining the mutation rate are often fossil or archaeological dates. The molecular clock was first tested in 1962 on the hemoglobin protein variants of various animals, and is commonly used in molecular evolution to estimate times of speciation or radiation. It is sometimes called a gene clock or an evolutionary clock. Early discovery and genetic equidistance The notion of the existence of a so-called "molecular clock" was first attributed to Émile Zuckerkandl and Linus Pauling who, in 1962, noticed that the number of amino acid differences in hemoglobin between different lineages changes roughly linearly with time, as estimated from fossil evidence. T

Circadian rhythm and cell cycle

Rosamaria Cannavo

Circadian rhythms are biological processes found in most living organisms, displaying a roughly 24-hour period, responding primarily to light darkness cycles in an organism's environment. At the cellular level, the circa-24h rhythmicity is generated by a molecular clock based on a transcription-translation feedback network and consists of a cell-autonomous and self-sustained oscillator. In conditions where cells proliferate, the cell division cycle can be also considered as an oscillator. Since both processes run with similar periods in several mammalian cells, it is reasonable to expect that interactions between these two cycles may cause synchronization. Many studies reported evidences of interactions between the circadian and the cell cycles in different organisms. In particular, it appeared that in several systems, specific cell cycle phases occur in distinct temporal windows rather than being randomly distributed in time. These findings led to the concept of circadian gating of the cell cycle, through which the circadian clock can favor or forbid cell cycle transitions at specific circadian phases. However, it was also reported the converse, namely an effect of cell division on the circadian oscillator. Even though interactions between the circadian clock and the cell cycle have been identified in both directions, the dynamical consequences and the directionality of the coupling at the single-cell level were not extensively investigated. In order to better characterize the potential synchronization in mammalian cells, we estimated the mutual interactions between circadian clock and cell cycle in NIH3T3 mouse fibroblasts by the use of time-lapse fluorescent microscopy in combination with statistical analysis and mathematical modeling. NIH3T3 cells, harboring a fluorescent reporter under the control of the circadian RevErb-a gene promoter, were imaged for several days allowing the simultaneous detection of circadian oscillations and time of divisions. The analysis of thousands of circadian cycles in dividing cells indicated that both oscillators are synchronized, with cell divisions occurring about 5 h before the peak of the circadian RevErb-a reporter. We tested several perturbations such as different serum concentrations, different temperatures, treatment with pharmacological compounds and shRNA-mediated knockdown of circadian regulators. Surprisingly, this showed that circadian rhythm and cell cycle remain synchronized over the wide range of conditions probed. Our data showed that this synchronization state reflects an unexpected predominant influence of the cell cycle on the circadian oscillator, and did not support the leading hypothesis about a circadian gating of the cell cycle. The stochastic modeling of two interacting phase oscillators allowed us to identify the parameters of the coupling functions, revealing an acceleration of circadian phase after the division. The work presented in this thesis sheds light on the interaction between two fundamentally recurrent cellular processes in mammalian cells and provides a deeper understanding of the role of the circadian clock in proliferating cells and tissues. These findings might have significant implications for chronobiology and chronotherapeutics.

EPFL2016

Phase specific transcriptional regulation of circadian clock and metabolism in mouse liver

Jonathan Aryeh Sobel

The molecular clock has been conserved from cyanobacteria to mammals and is believed to align behavioral and biochemical processes with the diurnal cycle. This cellular mechanism has been an advantage to increase the fitness of organisms through the ability to anticipate food availability or predator presence. Accumulating evidence has revealed that the circadian clock is intimately interconnected with the metabolic cycle. But, the nature of these interconnections is yet not clear at the transcriptional level, and the contribution of cis-regulatory modules has not been elucidated. Accessible chromatin regions of the genome are implicated in diverse processes, such as gene regulation through enhancers and promoters, insulation of genomic domains or alternative splicing. They allow cell-type specific programs that are controlled by tissue-specific transcription factors and chromatin modifiers, although tissue-specific regulation of the circadian clock remains unclear. Therefore, we compared genome-wide BMAL1 binding and chromatin accessibility in the liver and in NIH3T3 fibroblasts. Moreover, for the first time, our study explores the dynamics of accessible regions, histone 3 lysine 27 acetylation (H3K27ac) and RNA polymerase II (Pol II) every 4 hours over one day in mouse liver. In this study, we show that a substantial fraction of these accessible sites are oscillating during a diurnal cycle with a circadian period. We observed that these accessible regions are enriched in the proximity of actively transcribed genes, and that they are dynamically affected by the binding of transcription factors such as BMAL1. We investigated wild-type (WT) and Bmal1-/- genotypes, in night restricted feeding regimen and in Light-Dark (LD) cycle, to study the circadian clock regulatory network underlying diurnal transcription. Therefore, we applied a penalized generalized linear model to infer the activity of transcription factor binding motifs in oscillating accessible sites using Pol II loadings at the transcription start sites of nearby genes. We were able to recapitulate the known regulatory elements of the circadian clock, notably E-box, D-Box, and ROR-responsive elements (RRE) in the WT genotype. On the other hand, we found that Forkhead box (FOX), glucocorticoids responsive elements (GRE), and C-AMP Response Element (CRE) were the main contributors of the regulation of oscillating genes in Bmal1-/-. Finally, using a mixture model to detect footprints with a base pair resolution, we studied the dynamics of the accessibility overlapping E-box. Our last analysis suggested that BMAL1/CLOCK is binding on double E-boxes with a spacer of 6 or 7 bp in a hetero-tetramer configuration. A 3D structure model further supported this binding mode. In Summary, we used DNase I-seq and ChIP-seq of Pol II and H3K27ac to study the circadian chromatin landscape in a 4h time-resolved experimental design. We uncovered the underlying circadian transcriptional regulatory network and, we dissected the chromatin accessibility around BMAL1 binding sites at a base pair resolution, which led to an unappreciated mode of binding of BMAL1/CLOCK in a hetero-tetramer conformation on double E-boxes. Lastly, we found tissue-specific factors that might contribute to tissue-specific binding of BMAL1/CLOCK.

EPFL2016

Systems Chronobiology: Global Analysis of Gene Regulation in a 24-Hour Periodic World

Jérôme Mermet, Felix Naef, Jake Yeung

Mammals have evolved an internal timing system, the circadian clock, which synchronizes physiology and behavior to the daily light and dark cycles of the Earth. The master clock, located in the suprachiasmatic nucleus (SCN) of the brain, takes fluctuating light input from the retina and synchronizes other tissues to the same internal rhythm. The molecular clocks that drive these circadian rhythms are ticking in nearly all cells in the body. Efforts in systems chronobiology are now being directed at understanding, on a comprehensive scale, how the circadian clock controls different layers of gene regulation to provide robust timing cues at the cellular and tissue level. In this review, we introduce some basic concepts underlying periodicity of gene regulation, and then highlight recent genome-wide investigations on the propagation of rhythms across multiple regulatory layers in mammals, all the way from chromatin conformation to protein accumulation.

Cold Spring Harbor Lab Press, Publications Dept2017