Skene, D. J. | bioRxiv Science

bioRxiv2024

Rhythmic variation in proteomics: challenges and opportunities for statistical power and biomarker identification

Time-of-day variation in the molecular profile of biofluids and tissues is a well-described phenomenon, but - especially for proteomics - is rarely considered in terms of the challenges this presents to reproducible biomarker identification. In this work we demonstrate these confounding issues using a small-scale proteomics analysis of male participants in a constant routine protocol following an 8-day laboratory study, in which sleep-wake, light-dark and meal timings were controlled. We provide a case study analysis of circadian and ultradian rhythmicity in proteins in the complement and coagulation cascades, as well as apolipoproteins, and demonstrate that rhythmicity increases the risk of Type II errors due to the reduction in statistical power from increased variance. For the proteins analysed herein we show that to maintain statistical power if chronobiological variation is not controlled for, n should be increased (by between 9% and 20%); failure to do so would increase {beta}, the chance of Type II error, from a baseline value of 20% to between 22% and 28%. Conversely, controlling for rhythmic time-of-day variation in study design offers the opportunity to improve statistical power and reduce the chances of Type II errors. Indeed, control of time-of-day sampling is a more cost-effective strategy than increasing sample sizes. We recommend that best practice in proteomics study design should account for temporal variation as part of sampling strategy where possible. Where this is impractical, we recommend that additional variance from chronobiological effects be considered in power calculations, that time of sampling be reported as part of study metadata, and that researchers reference any previously identified rhythmicity in biomarkers and pathways of interest. These measures would mitigate against both false and missed discoveries, and improve reproducibility, especially in studies looking at biomarkers, pathways or conditions with a known chronobiological component.

molecular biology↗

bioRxiv2020

An essential amino acid synchronises malaria parasite development with daily host rhythms

Rapid asexual replication of blood stage malaria parasites is responsible for the severity of disease symptoms and fuels the production of transmission forms. That malaria parasite species coordinate their cycles of asexual replication with daily rhythms of their host was discovered in the Hippocratic era, but how and why this occurs is enigmatic. Here, we demonstrate that the Plasmodium chabaudis schedule for asexual replication can be orchestrated by a isoleucine, metabolite provided to the parasite in periodic manner due to the hosts rhythmic intake of food. First, we identify nutrients with daily rhythms in the blood that match the timing of rhythms in both host feeding and the developmental schedule of asexually replicating parasites. We hypothesise that if parasites set their own developmental schedule, they should use a time-of-day cue that is a factor they cannot generate endogenously at any time-of-day, or scavenge in a round-the-clock manner. Our large-scale metabolomics experiment reveals that only one metabolite - the amino acid isoleucine - fits these criteria. Second, further experiments reveal that parasites alter the developmental schedule of asexual stages in response to isoleucine provision and withdrawal in the manner consistent with it acting as a time-cue. Specifically, parasites respond to isoleucine loss by slowing development. This is a parasite strategy rather than the consequences of an imposed constraint, because unlike when parasites are deprived of other essential nutrients, they suffer no apparent costs in the absence of isoleucine. Overall, our data suggest parasites can use the daily rhythmicity of blood-isoleucine concentration to synchronise asexual development with the availability of isoleucine, and potentially other resources, that arrive in the blood in a periodic manner due to the hosts daily feeding-fasting cycle. Identifying both how and why parasites keep time opens avenues for interventions; interfering with the parasites time-keeping mechanism may stall replication, increasing the efficacy of drugs and immune responses, and could also prevent parasites from entering dormancy to tolerate drugs.

microbiology↗