Extended Data Figure 9. The effects of magnesium depletion and role of mTOR.

a. Extracellular lactate was measured in U2OS cells after 24 hours in Mg²⁺-depleted compared with normal media. b,c. Combined action of extracellular magnesium depletion and mTOR inhibition using torin1 (b, n=3) or rapamycin (c, n=6) to lengthen circadian period in U2OS cells is less than additive (mean±SEM). d,e. Quantification of period lengthening due to torin1 (d, n=3) and rapamycin (e, n=6) in Mg²⁺-depleted compared with normal media (mean±SEM). Note the apparent ‘ceiling effect’ at high concentrations of both drugs, such that Mg²⁺-depletion elicits no additional lengthening of cellular circadian period. Two-way ANOVA interaction effect: p<0.0001 for both drugs vs. Mg²⁺, selected p-values for Sidak’s multiple comparisons test are also reported (n.s., p>0.33).