Approaches to Engineer Stability of Beetle Luciferases

Koksharov, M. I.; Ugarova, N. N.

doi:10.5936/csbj.201209004

Cited by 18 publications

(13 citation statements)

References 68 publications

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“…6a). Since temperature impacts the enzymatic activity of the luciferase 27 , we first corrected the signal for this systematic effect (Supplementary Information). We found that, independently of initial densities, as the populations reach confluency (which takes longer for initial low density), the phases and amplitudes become stationary, showing 1:1 entrainment (Fig.…”

Section: Resultsmentioning

confidence: 99%

Low-dimensional Dynamics of Two Coupled Biological Oscillators

Droin

Paquet

Naef

2019

Preprint

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The circadian clock and the cell cycle are two biological oscillatory processes that coexist within individual cells. These two oscillators were found to interact, which can lead to their synchronization. Here, we develop a method to infer their coupling and non-linear dynamics from thousands of mouse and human single-cell microscopy traces. This coupling predicts multiple phase-locked states showing different degrees of robustness against molecular fluctuations inherent to cellular scale biological oscillators. Moreover, the phase–locked states were temperature-independent and evolutionarily conserved from mouse to human, hinting at a common underlying dynamical mechanism. Finally, we detected a signature of the coupled dynamics in a physiological context, where tissues with different proliferation states exhibited shifted circadian clock phases.

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Section: Resultsmentioning

confidence: 99%

Low-dimensional Dynamics of Two Coupled Biological Oscillators

Droin

Paquet

Naef

2019

Preprint

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“…The activity of the commonly used Photinus phyralis firefly luciferase is considerably higher at reduced temperatures. This effect is attributed to reduced protein activity and a so-called red shift in the luminescence at higher temperatures (for review, see Koksharov and Ugarova 2012). Therefore, we used the more temperature-tolerant mutant Luciola mingrelica firefly luciferase (LMLucR) (Koksharov and Ugarova 2011) for the construction of the Cirbp-luciferase reporter gene.…”

Section: Temperature-dependent Cirbp Mrna Accumulation Is Regulated Bmentioning

confidence: 99%

Temperature regulates splicing efficiency of the cold-inducible RNA-binding protein gene Cirbp

et al. 2016

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In mammals, body temperature fluctuates diurnally around a mean value of 36°C-37°C. Despite the small differences between minimal and maximal values, body temperature rhythms can drive robust cycles in gene expression in cultured cells and, likely, animals. Here we studied the mechanisms responsible for the temperaturedependent expression of cold-inducible RNA-binding protein (CIRBP). In NIH3T3 fibroblasts exposed to simulated mouse body temperature cycles, Cirbp mRNA oscillates about threefold in abundance, as it does in mouse livers. This daily mRNA accumulation cycle is directly controlled by temperature oscillations and does not depend on the cells' circadian clocks. Here we show that the temperature-dependent accumulation of Cirbp mRNA is controlled primarily by the regulation of splicing efficiency, defined as the fraction of Cirbp pre-mRNA processed into mature mRNA. As revealed by genome-wide "approach to steady-state" kinetics, this post-transcriptional mechanism is widespread in the temperature-dependent control of gene expression.[Keywords: Cirbp; splicing efficiency; temperature; circadian rhythms] Supplemental material is available for this article. Temperature is a fundamental physical parameter that influences metabolism in all organisms by modulating the rate of biochemical reactions. Endothermic organisms like mammals have acquired special thermoregulatory adaptation mechanisms in order to decrease their dependence on environmental temperature. These mechanisms maintain core body temperature (CBT) within a favorable range, allowing efficient metabolic activity throughout diurnal and seasonal cycles (for review, see Tattersall 2012).Although regulated around a species-specific set point, mammalian CBT shows diurnal oscillations of ∼0.4°C-6.0°C (for review, see Refinetti 2010). These regular oscillations are generated under the control of the circadian timekeeping system, which is composed of a master pacemaker in the brain's suprachiasmatic nucleus (SCN) and subsidiary peripheral clocks in nearly all other cells of the body (Dibner et al. 2010). Although peripheral circadian oscillators are self-sustained and cell-autonomous, they must be synchronized by the SCN and/or environmental Zeitgebers (timing cues), such as light-dark cycles, in order to maintain phase coherence and rhythmic physiology in the body. Without a functional SCN, laboratory rodents kept in constant darkness lose virtually all overt rhythms in behavior and physiology, including CBT cycles (Refinetti et al. 1994).While the period length (τ) of circadian oscillations is remarkably resilient to temperature variations, a phenomenon called temperature compensation, the phase is exquisitely sensitive to temperature perturbations, in particular in peripheral cells. In fact, in cultured fibroblasts and tissue explants, the phase of circadian clock gene expression can be perfectly synchronized by simulated CBT cycles (Brown et al. 2002;Buhr et al. 2010;Saini et al. 2012).In intact animals, the systemic synchronization cues governed by t...

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“…The design of proteins with enhanced thermostability is one of the major goals of protein engineering. Many attempts have been made by site directed mutagenesis to increase thermostability of firefly luciferase [3][4][5].…”

Section: Introductionmentioning

confidence: 99%