Transient systemic autophagy ablation irreversibly inhibits lung tumor cell metabolism and promotes T-cell mediated tumor killing.

Macroautophagy/autophagy is a highly conserved catabolic process pivotal to cellular homeostasis and support of tumorigenesis. Being a potential therapeutic target for cancer, we have worked to understand the implications of autophagy inhibition both systemically, and tumor-specifically. We utilized inducible expression of Atg5 shRNA to temporally control autophagy levels in a reversible manner to study the effects of tumor-intrinsic and systemic autophagic loss and restoration on established KrasG12D/+;trp53-/- (KP) lung tumor growth. We reported that transient systemic ATG5 loss significantly reduces KP lung tumor growth. Through in vivo isotope tracing and metabolic flux analyses, we noted that systemic ATG5 knockdown significantly reduces the uptake of glucose and lactate in lung tumors, leading to impaired TCA cycle metabolism and biosynthesis. Additionally, we observed an increased tumor T cell infiltration in the absence of systemic ATG5, which is essential for T cell-mediated tumor killing. Moreover, the impaired tumor metabolism and increased T cell infiltration are sustained when autophagy is restored in a short term. Finally, we found that intermittent systemic ATG5 knockdown, a mock therapy situation, significantly prolongs the lifespan of mice bearing KP lung tumors. Our findings lay the proof of concept for inhibition of autophagy as a valid approach to cancer therapy.

Structural insight into transmissive mutant huntingtin species by correlative light and electron microscopy and cryo-electron tomography.

Aggregates of mutant huntingtin (mHTT) containing an expanded polyglutamine (polyQ) tract are hallmarks of Huntington's Disease (HD). Studies have shown that mHTT can spread between cells, leading to the propagation of misfolded protein pathology. However, the structure of transmissive mHTT species, and the molecular mechanisms underlying their transmission remain unknown. Using correlative light and electron microscopy (CLEM) and cryo-electron tomography (cryo-ET), we identified two types of aggregation-prone granules in conditioned medium from PC12 cells expressing a mHTT N-terminal fragment: densities enclosed by extracellular vesicles (EVs), and uncoated, amorphous meshworks of heterogeneous oligomers that co-localize with clusters of EVs. In vitro assays confirmed that liposomes induce condensation of polyQ oligomers into higher-order assemblies, resembling the uncoated meshworks observed in PC12 conditioned medium. Our findings provide novel insights into formation and architecture of transmissive mHTT proteins, and highlight the potential role of EVs as both carriers and modulators of transmissive mHTT proteins.