Preprint: mTOR inhibition eliminated most daily fluctuations in mouse liver

Preprint: mTOR inhibition eliminated most daily fluctuations in mouse liver

In a preprint published on August 31, researchers temporarily inhibited mTOR in cells and mice. Many daily changes disappeared from the cells and tissues, although the proteins that keep the biological clock running continued to oscillate. In the liver, 86% of the 639 proteins whose abundance varied over the course of a day lost this variation.

The circadian clock consists of proteins and genes that maintain an approximately 24-hour rhythm within a cell. The conventional model links the clock to rhythmic gene activity. However, RNA abundance often changes differently from the abundance of the corresponding proteins, and most proteins persist for more than a day. The authors investigated a mechanism through which the clock might regulate cellular processes: mTOR, a signaling system that responds to nutrients, energy reserves, and growth signals and regulates protein synthesis and metabolism.

In mouse fibroblasts, which are connective tissue cells, the authors used INK128 to inhibit both mTOR complexes. A luminescent reporter that reflected the abundance of the clock protein PER2 continued to oscillate, but the oscillations became weaker and their period lengthened by approximately 10%. At the same time, daily changes disappeared in protein synthesis and degradation, respiration, nuclear RNA abundance, and the rate at which an experimental wound closed in a cell layer. This allowed the authors to distinguish the preserved clock rhythm from the cellular processes that it normally coordinates.

After three days of partial mTOR inhibition in mouse liver, 639 proteins varied with the time of day. These changes disappeared in 86% of them. The rhythms of CRY1 and PER1, two clock proteins, persisted. Daily changes in RNA, gas exchange, and hydroxybupropion levels also weakened. Hydroxybupropion is produced during the metabolism of bupropion. In the forebrain, most daily changes in protein abundance and phosphorylation disappeared. Phosphorylation adds chemical modifications that alter protein activity. The mice largely retained their daily sleep and wake pattern, but they remained awake longer around transitions between light and darkness.

In human cells, PER2 physically interacted with mTORC1, one of the mTOR complexes. Rapid removal of PER2 increased the phosphorylation of mTORC1 target proteins. The authors propose a model in which PER2 oscillations tune mTOR activity, while mTOR conveys the timing signal to cellular processes.

In a plant and a fungus, inhibition of TOR, a protein related to mTOR, weakened the rhythms of physiological processes more strongly than it affected the biological clock itself. This finding is consistent with the authors' proposal that TOR transmits signals from the biological clock to physiological processes across very different groups of organisms.