Retromer prevents the destruction of numerous receptors by recycling them from endosomes to the trans-Golgi network or plasma membrane. This enables retromer to fine-tune the activity of many signaling pathways in parallel. However, the mechanism(s) by which retromer function adapts to environmental fluctuations such as nutrient withdrawal and how this affects the fate of its cargoes remains incompletely understood. Here, we reveal that macroautophagy/autophagy inhibition by MTORC1 controls the abundance of retromer⁺ endosomes under nutrient-replete conditions. Autophagy activation by chemical inhibition of MTOR or nutrient withdrawal does not affect retromer assembly or its interaction with the RAB7 GAP protein TBC1D5, but rather targets these endosomes for bulk destruction following their capture by phagophores. This process appears to be distinct from amphisome formation. TBC1D5 and its ability to bind to retromer, but not its C-terminal LC3-interacting region (LIR) or nutrient-regulated dephosphorylation, is critical for retromer to be captured by autophagosomes following MTOR inhibition. Consequently, endosomal recycling of its cargoes to the plasma membrane and trans-Golgi network is impaired, leading to their lysosomal turnover. These findings demonstrate a mechanistic link connecting nutrient abundance to receptor homeostasis.

Autophagy slows age‐related pathologies and is stimulated by nutrient restriction in animal studies. However, this has never been shown in humans. We measured autophagy using a physiologically relevant measure of autophagic flux (flux of MAP1LC3B isoform II/LC3B‐II in peripheral blood mononuclear cells in the context of whole blood) in 121 humans with obesity who were randomised to standard care (SC, control condition), calorie restriction (CR) or intermittent fasting plus time‐restricted eating (iTRE) for 6 months. While the differences in change from baseline between groups was not significant at 2 months, we observed a significant difference in change from baseline between iTRE compared to SC at 6 months (P = 0.04, post hoc analysis). This effect may be driven partly by a tendency for autophagy to decrease in the SC group. The difference in change from baseline between CR and SC was not significant. Uncorrected analysis of correlations showed a negative relationship between change in autophagy and change in blood triglycerides. Data on the specificity and performance of the methods used to measure human autophagy are also presented. This shows autophagy may be increased by intermittent nutrient restriction in humans. If so, this is a demonstration that nutrient restriction can be used to improve a primary hallmark of biological ageing and provides a mechanism for how fasting could delay the onset of age‐related disease. image Key points: Autophagy slows biological ageing, and dysfunction of autophagy has been implicated in age‐related disease – an effective way of increasing autophagy in cells and animal models is nutrient restriction. However, the impact of different types of nutrient restriction on physiological autophagic flux in humans has not been extensively researched. Here we measure the effect of intermittent time‐restricted eating (iTRE) and calorie restriction on physiological autophagic flux in peripheral blood mononuclear cells. After 6 months, there was a significant difference in change from baseline between the iTRE and the standard care control group, with flux being higher in the iTRE group at this timepoint. However, there was no significant increase from baseline within the iTRE group, showing that although autophagy may be modified by nutrient restriction in humans, further studies are required.

Autophagy is a ‘waste-disposal’ pathway that protects against age-related pathology. It is widely accepted that autophagy declines with age, yet the role that sex and diet-related obesity play during aging remain unknown. Here, we present the most comprehensive in vivo study of autophagic flux to date. We employed transgenic mice overexpressing tandem-florescent LC3B (RFP-GFP-LC3B) to measure autophagic flux in the blood (PBMCs), heart, and motor cortex neurons of aging mice that were fed regular chow or a high-fat diet for 6-, 12- or 18-months. In male mice, aging decreased autophagic flux in the heart, increased it in the blood, and had no effect in motor cortex neurons. Age-dependent changes autophagic flux were less pronounced in female mice. High-fat diet influenced autophagic flux in the blood and heart of male but not female mice. Overall, we uncovered sexual dimorphisms that underpin how autophagy changes with age across different tissues and in response to a high-fat diet.