Ras-Mediated Deregulation of the Circadian Clock in Cancer

Abstract

Circadian rhythms are essential to the temporal regulation of molecular processes in living systems and as such to life itself. Deregulation of these rhythms leads to failures in biological processes and eventually to the manifestation of pathological phenotypes including cancer. To address the questions as to what are the elicitors of a disrupted clock in cancer, we applied a systems biology approach to correlate experimental, bioinformatics and modelling data from several cell line models for colorectal and skin cancer. We found strong and weak circadian oscillators within the same type of cancer and identified a set of genes, which allows the discrimination between the two oscillator-types. Among those genes are IFNGR2, PITX2, RFWD2, PPARγ, LOXL2, Rab6 and SPARC, all involved in cancer-related pathways. Using a bioinformatics approach, we extended the core-clock network and present its interconnection to the discriminative set of genes. Interestingly, such gene signatures link the clock to oncogenic pathways like the RAS/MAPK pathway. To investigate the potential impact of the RAS/MAPK pathway - a major driver of colorectal carcinogenesis - on the circadian clock, we used a computational model which predicted that perturbation of BMAL1-mediated transcription can generate the circadian phenotypes similar to those observed in metastatic cell lines. Using an inducible RAS expression system, we show that overexpression of RAS disrupts the circadian clock and leads to an increase of the circadian period while RAS inhibition causes a shortening of period length, as predicted by our mathematical simulations. Together, our data demonstrate that perturbations induced by a single oncogene are sufficient to deregulate the mammalian circadian clock.

Author Summary

Living systems possess an endogenous time-generating system – the circadian clock - accountable for a 24 hours oscillation in the expression of about 10% of all genes. In mammals, disruption of oscillations is associated to several diseases including cancer. In this manuscript, we address the following question: what are the elicitors of a disrupted clock in cancer? We applied a systems biology approach to correlate experimental, bioinformatics and modelling data and could thereby identify key genes which discriminate strong and weak oscillators among cancer cell lines. Most of the discriminative genes play important roles in cell cycle regulation, DNA repair, immune system and metabolism and are involved in oncogenic pathways such as the RAS/MAPK. To investigate the potential impact of the Ras oncogene in the circadian clock we generated experimental models harbouring conditionally active Ras oncogenes. We put forward a direct correlation between the perturbation of Ras oncogene and an effect in the expression of clock genes, found by means of mathematical simulations and validated experimentally. Our study shows that perturbations of a single oncogene are sufficient to deregulate the mammalian circadian clock and opens new ways in which the circadian clock can influence disease and possibly play a role in therapy.