Macroautophagy is a highly conserved mechanism of lysosomal-mediated protein degradation that plays a key role in maintaining cellular homeostasis by recycling amino acids, reducing the amount of damaged proteins, and regulating protein levels in response to extracellular signals. We have found that macroautophagy is induced after effector T cell activation. Engagement of the TCR and CD28 results in enhanced microtubule-associated protein 1 light chain 3 (LC3) processing, increased numbers of LC3-containing vesicles, and increased LC3 flux, indicating active autophagosome formation and clearance. The autophagosomes formed in stimulated T cells actively fuse with lysosomes to degrade their cargo. Using a conditional KO mouse model where Atg7, a critical gene for macroautophagy, is specifically deleted in T cells, we have found that macroautophagy-deficient effector Th cells have defective IL-2 and IFN-γ production and reduced proliferation after stimulation, with no significant increase in apoptosis. We have found that ATP generation is decreased when autophagy is blocked, and defects in activation-induced cytokine production are restored when an exogenous energy source is added to macroautophagy-deficient T cells. Furthermore, we present evidence showing that the nature of the cargo inside autophagic vesicles found in resting T cells differs from the cargo of autophagosomes in activated T cells, where mitochondria and other organelles are selectively excluded. These results suggest that macroautophagy is an actively regulated process in T cells that can be induced in response to TCR engagement to accommodate the bioenergetic requirements of activated T cells.

Chaperone-mediated autophagy selectively targets single cytoplasmic proteins for degradation. Macian and colleagues show that such autophagy is induced by engagement of the TCR to target negative regulators of T cell activation. Chaperone-mediated autophagy (CMA) targets soluble proteins for lysosomal degradation. Here we found that CMA was activated in T cells in response to engagement of the T cell antigen receptor (TCR), which induced expression of the CMA-related lysosomal receptor LAMP-2A. In activated T cells, CMA targeted the ubiquitin ligase Itch and the calcineurin inhibitor RCAN1 for degradation to maintain activation-induced responses. Consequently, deletion of the gene encoding LAMP-2A in T cells caused deficient in vivo responses to immunization or infection with Listeria monocytogenes. Impaired CMA activity also occurred in T cells with age, which negatively affected their function. Restoration of LAMP-2A in T cells from old mice resulted in enhancement of activation-induced responses. Our findings define a role for CMA in regulating T cell activation through the targeted degradation of negative regulators of T cell activation.

Significance: Glioblastoma (GB) is the most lethal brain malignancy without an effective treatment. In this study, we demonstrate that tumor-induced change in chaperone-mediated autophagy (CMA) in host perivascular cells is a targetable process to prevent GB progression. CMA regulates pericyte interaction with tumor cells and sustains the acquired immunosuppressive function of pericytes, which is required for tumor survival. Blockage of CMA results in changes in the protein levels involved in cell-to-cell interaction and affects the pericyte secretory phenotype, resulting in defective GB adhesion and diminished tumor survival. This work reveals a previously unknown capacity of GB to modulate host pericyte CMA to assist in its own progression. Our results highlight the possibility of targeting CMA to treat this aggressive disease.

Glioblastoma progression is assisted by induction of immunosuppressive function of pericytes through interaction with tumor cells Glioblastoma progression is assisted by induction of immunosuppressive function of pericytes through interaction with tumor cells

A hallmark of aging is an imbalance between production and clearance of reactive oxygen species and increased levels of oxidatively damaged biomolecules. Herein, we demonstrate that splenic and nodal antigen-presenting cells purified from aging mice accumulate oxidatively modified proteins with side-chain carbonylation, advanced glycation end products, and lipid peroxidation. Furthermore, we show that the endosomal accumulation of oxidatively modified proteins interferes with the efficient processing of exogenous antigens and degradation of macroautophagy-delivered proteins. In support of a causative role for oxidized products in the inefficient immune response, a decrease in oxidative stress improved the adaptive immune response to immunizing antigens. These findings underscore a previously unrecognized negative effect of age-dependent changes in cellular proteostasis on the immune response.