The mammalian target of rapamycin (mTOR) pathway, which is essential for cell proliferation, is repressed in certain cell types in hypoxia. However, hypoxia-inducible factor 2α (HIF2α) can act as a proliferation-promoting factor in some biological settings. This paradoxical situation led us to study whether HIF2α has a specific effect on mTORC1 regulation. Here we show that activation of the HIF2α pathway increases mTORC1 activity by upregulating expression of the amino acid carrier SLC7A5. At the molecular level we also show that HIF2α binds to the Slc7a5 proximal promoter. Our findings identify a link between the oxygen-sensing HIF2α pathway and mTORC1 regulation, revealing the molecular basis of the tumor-promoting properties of HIF2α in von Hippel-Lindau-deficient cells. We also describe relevant physiological scenarios, including those that occur in liver and lung tissue, wherein HIF2α or low-oxygen tension drive mTORC1 activity and SLC7A5 expression.

Variations in background 2H and 18O abundances in body water influence the accuracy and precision of the 2H218O method for determination of energy expenditure. To investigate the effect of weaning during infancy on background 2H and 18O abundances, urine samples from 44 breast- or formula-fed infants aged 5-16 wk were analyzed. 2H and 18O abundances were significantly higher (P less than 0.001) in breast- than in formula-fed infants. The relationship between 2H and 18O abundances was linear and independent of diet [slope, 4.16 +/- 0.43 (SE)]. By use of this information, the effect of weaning on the accuracy of the 2H218O method was evaluated, taking into account the effect of 2H-18O abundances in the isotope loading dose. In infants weaned completely from breast milk to formula during the measurement, energy expenditure can be overestimated by 18.0%, even if 2H-18O abundances in the isotope dose equal the ratio of naturally occurring background changes. However, this error can be reduced to less than 3.0% by manipulating the study duration and isotope dose. During gradual weaning, the overestimation of energy expenditure is only 0.3-2.0%.

The doubly labeled water method was compared with indirect calorimetry and a nutrient-balance study for simultaneous determination of rates of CO2 production, energy expenditure, and water intake over 5 days in four preterm infants. Additionally, metabolizable energy (ME) intake estimated using the isotope procedure (as energy expenditure plus an estimate for energy deposition based on weight gain), was compared to ME intake measured in the balance study. Compared to values obtained by traditional methods, calculated CO2 production, energy expenditure, and water intake differed by -1.4 +/- 4.8% (SD), +0.3 +/- 2.6%, and +5.7 +/- 1.4%, respectively; the difference in water intake was significant (p less than 0.05). Calculated ME intakes were 5.3 +/- 19.3% less than measured intakes, but the difference was not significant. These findings indicate that the doubly labeled water method can provide accurate information on rates of CO2 production, energy expenditure, and water intake in preterm infants, but individual estimates of ME intake may be subject to substantial error.

The composition and physical properties of the extracellular matrix (ECM) critically influence tumor progression, but the molecular mechanisms underlying ECM layering are poorly understood. Tumor–stroma interaction critically depends on cell communication mediated by exosomes, small vesicles generated within multivesicular bodies (MVBs). We show that caveolin-1 (Cav1) centrally regulates exosome biogenesis and exosomal protein cargo sorting through the control of cholesterol content at the endosomal compartment/MVBs. Quantitative proteomics profiling revealed that Cav1 is required for exosomal sorting of ECM protein cargo subsets, including Tenascin-C (TnC), and for fibroblast-derived exosomes to efficiently deposit ECM and promote tumor invasion. Cav1-driven exosomal ECM deposition not only promotes local stromal remodeling but also the generation of distant ECM-enriched stromal niches in vivo. Cav1 acts as a cholesterol rheostat in MVBs, determining sorting of ECM components into specific exosome pools and thus ECM deposition. This supports a model by which Cav1 is a central regulatory hub for tumor–stroma interactions through a novel exosome-dependent ECM deposition mechanism.