Circadian Analysis of the Mouse Cerebellum Proteome.

Marine Plumel,Cristina Sandu,Fabrice Bertile,Marie-Paule Felder-Schmittbuhl,Pauline Maes,Etienne Challet,Stéphanie Dumont

doi:10.3390/ijms20081852

Abstract

The cerebellum contains a circadian clock, generating internal temporal signals. The daily oscillations of cerebellar proteins were investigated in mice using a large-scale two-dimensional difference in gel electrophoresis (2D-DIGE). Analysis of 2D-DIGE gels highlighted the rhythmic variation in the intensity of 27/588 protein spots (5%) over 24 h based on cosinor regression. Notably, the rhythmic expression of most abundant cerebellar proteins was clustered in two main phases (i.e., midday and midnight), leading to bimodal distribution. Only six proteins identified here to be rhythmic in the cerebellum are also known to oscillate in the suprachiasmatic nuclei, including two proteins involved in the synapse activity (Synapsin 2 [SYN2] and vesicle-fusing ATPase [NSF]), two others participating in carbohydrate metabolism (triosephosphate isomerase (TPI1] and alpha-enolase [ENO1]), Glutamine synthetase (GLUL), as well as Tubulin alpha (TUBA4A). Most oscillating cerebellar proteins were not previously identified in circadian proteomic analyses of any tissue. Strikingly, the daily accumulation of mitochondrial proteins was clustered to the mid-resting phase, as previously observed for distinct mitochondrial proteins in the liver. Moreover, a number of rhythmic proteins, such as SYN2, NSF and TPI1, were associated with non-rhythmic mRNAs, indicating widespread post-transcriptional control in cerebellar oscillations. Thus, this study highlights extensive rhythmic aspects of the cerebellar proteome.

Highlights

The circadian system controls most aspects of behavior and metabolism on a 24 h time-scale
The molecular mechanisms of the circadian clocks rely on auto-regulatory transcriptional and translational feedback loops that generate a rhythmic regulation of specific genes, called clock genes
This led to the detection of 588 protein spots, among which 26 (i.e., 5% from all spots) were found to be rhythmic according to cosinor procedure, with an additional one being very close to significance

Summary

Introduction

The circadian system controls most aspects of behavior and metabolism on a 24 h time-scale. Circadian rhythmicity regulates the timing of physiological mechanisms via daily organisation (e.g., anticipation of predictable events) and partitioning (e.g., temporal segregation of incompatible functions) at the cellular, tissue and organism levels. The molecular mechanisms of the circadian clocks rely on auto-regulatory transcriptional and translational feedback loops that generate a rhythmic regulation of specific genes, called clock genes. A negative loop involves PER and CRY proteins that repress transactivation mediated by CLOCK-BMAL1 [3]. A number of clock proteins undergo post-translational modifications, such as phosphorylation, acetylation, ubiquitination, and sumoylation that modulate characteristics (i.e., phase, amplitude or period) of circadian oscillations [7,8]. The positive circadian loop mediated by CLOCK-BMAL1 dimers controls the rhythmic transcription of clock-controlled genes, whose proteins provide temporal cues within and eventually out of the clock cells [3]

Methods

Results

Conclusion