Assessment of Mitochondrial Biogenesis and mTORC1 Signaling During Chronic Rapamycin Feeding in Male and Female Mice

Abstract

Chronic inhibition of the protein synthesis regulator mTORC1 through rapamycin extends life span in mice, with longer extension in females than in males. Whether rapamycin treatment inhibits protein synthesis or whether it does so differently between sexes has not been examined. UM-HET3 mice were fed a control or rapamycin-supplemented (Rap) diet for 12 weeks. Protein synthesis in mixed, cytosolic (cyto), and mitochondrial (mito) fractions and DNA synthesis and mTORC1 signaling were determined in skeletal muscle, heart, and liver. In both sexes, mito protein synthesis was maintained in skeletal muscle from Rap despite decreases in mixed and cyto fractions, DNA synthesis, and rpS6 phosphorylation. In the heart, no change in protein synthesis occurred despite the decreased DNA synthesis. In the heart and liver, Rap males were more sensitive to mTORC1 inhibition than Rap females. In conclusion, we show changes in protein synthesis and mTORC1 signaling that differ by sex and tissue.

Reduced in vivo hepatic proteome replacement rates but not cell proliferation rates predict maximum lifespan extension in mice

“…1C). This lack of any significant effect of Rapa treatment on the proliferation rates of any of these cell types contrasts with reports that Rapa reduces the proliferation rates of HepG2 liver cells in vitro and muscle cells in vivo (Varma & Khandelwal, 2007; Drake et al ., 2013). Divergence from these previously published results may be explained by differences in the dose and method of Rapa administration, differences in the in vitro versus the in vivo effects of Rapa, or differences in cell types.…”

“…Divergence from these previously published results may be explained by differences in the dose and method of Rapa administration, differences in the in vitro versus the in vivo effects of Rapa, or differences in cell types. However, our observation that the in vivo proliferation rate of liver cells was unchanged is consistent with a previous study that used a very similar experimental design (Drake et al ., 2013). …”

Assessment of protein synthesis in highly aerobic canine species at the onset and during exercise training

“…Mitochondrial biogenesis is, therefore, beneficial to sustain high aerobic energy production, but is also reliant on adequate energy availability. In accordance with this idea, we have previously demonstrated in rodent models that during energetic stress, associated with an increase in AMP-activated protein kinase (AMPK) activity or a decrease in the activity of the mechanistic target of rapamycin (mTOR), mitochondrial proteins are often selectively translated (5,10). This selective translation of mitochondrial proteins could be a countermeasure to energetic restriction.…”

“…Skeletal muscle tissue was fractionated according to our previously published procedures (4,5,13,14). Muscle (15-50 mg) was homogenized 1:10 with a bead homogenizer (Next Advance, Averill Park, NY) in isolation buffer (100 mM KCl, 40 mM Tris HCl, 10 mM Tris Base, 5 mM MgCl2, 1 mM EDTA, 1 mM ATP, pHϭ7.5) with phosphatase and protease inhibitors (HALT, Thermo Scientific, Rockford, IL).…”

“…Longer periods of assessment are limited by the need to maintain a constant infusion of isotopes, thus restricting freeliving conditions. To address these limitations, we (4,5,13,14,19,22) and others (2,6,16,26) have employed stable isotope techniques that use deuterium oxide ( 2 H 2 O). By providing 2 H 2 O in the drinking water of humans (19,22) or rodents (4,5,13,14), we have assessed cumulative protein synthesis over the period of weeks, rather than hours.…”