WIPIng the Brakes off Autophagy Induction.

In this issue of Molecular Cell, Wan et al. (2018) uncover WIPI2 as a critical rheostat in the control of autophagic degradation by mTORC1 and demonstrate the physiological utility of this signaling axis in promoting the clearance of hepatic lipids.

Autophagy in PDGFRα+ mesenchymal cells is essential for intestinal stem cell survival.

Autophagy defects are a risk factor for inflammatory bowel diseases (IBDs) through unknown mechanisms. Whole-body conditional deletion of autophagy-related gene (Atg) Atg7 in adult mice (Atg7Δ/Δ) causes tissue damage and death within 3 mo due to neurodegeneration without substantial effect on intestine. In contrast, we report here that whole-body conditional deletion of other essential Atg genes Atg5 or Fip200/Atg17 in adult mice (Atg5Δ/Δ or Fip200Δ/Δ) caused death within 5 d due to rapid autophagy inhibition, elimination of ileum stem cells, and loss of barrier function. Atg5Δ/Δ mice lost PDGFRα+ mesenchymal cells (PMCs) and Wnt signaling essential for stem cell renewal, which were partially rescued by exogenous Wnt. Matrix-assisted laser desorption ionization coupled to mass spectrometry imaging (MALDI-MSI) of Atg5Δ/Δ ileum revealed depletion of aspartate and nucleotides, consistent with metabolic insufficiency underlying PMC loss. The difference in the autophagy gene knockout phenotypes is likely due to distinct kinetics of autophagy loss, as deletion of Atg5 more gradually extended lifespan phenocopying deletion of Atg7 or Atg12. Thus, autophagy is required for PMC metabolism and ileum stem cell and mammalian survival. Failure to maintain PMCs through autophagy may therefore contribute to IBD.

The pleiotropic functions of autophagy in metastasis.

Autophagy is deregulated in many cancers and represents an attractive target for therapeutic intervention. However, the precise contributions of autophagy to metastatic progression, the principle cause of cancer-related mortality, is only now being uncovered. While autophagy promotes primary tumor growth, metabolic adaptation and resistance to therapy, recent studies have unexpectedly revealed that autophagy suppresses the proliferative outgrowth of disseminated tumor cells into overt and lethal macrometastases. These studies suggest autophagy plays unexpected and complex roles in the initiation and progression of metastases, which will undoubtedly impact therapeutic approaches for cancer treatment. Here, we discuss the intricacies of autophagy in metastatic progression, highlighting and integrating the pleiotropic roles of autophagy on diverse cell biological processes involved in metastasis.

Fibroblasts as architects of cancer pathogenesis.

Studies of epithelial cancers (i.e., carcinomas) traditionally focused on transformation of the epithelium (i.e., the cancer cells) and how aberrant signaling within the cancer cells modulates the surrounding tissue of origin. In more recent decades, the normal cells, blood vessels, molecules, and extracellular components that surround the tumor cells, collectively known as the "tumor microenvironment" or "stroma", have received increasing attention and are now thought to be key regulators of tumor initiation and progression. Of particular relevance to the work reviewed herein are the fibroblasts, which make up the major cell type within the microenvironment of most carcinomas. Due to their inherent heterogeneity, plasticity, and function, it is perhaps not surprising that fibroblasts are ideal modulators of normal and cancerous epithelium; however, these aspects also present challenges if we are to interrupt their tumor-supportive functions. Here, we review the current body of knowledge and the many questions that still remain about the special entity known as the cancer-associated fibroblast. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease.

Secretory autophagy maintains proteostasis upon lysosome inhibition.

The endolysosome system plays central roles in both autophagic degradation and secretory pathways, including the release of extracellular vesicles and particles (EVPs). Although previous work reveals important interconnections between autophagy and EVP-mediated secretion, our understanding of these secretory events during endolysosome inhibition remains incomplete. Here, we delineate a secretory autophagy pathway upregulated in response to endolysosomal inhibition, which mediates EVP-associated release of autophagic cargo receptors, including p62/SQSTM1. This secretion is highly regulated and dependent on multiple ATGs required for autophagosome formation, as well as the small GTPase Rab27a. Furthermore, disrupting autophagosome maturation, either via genetic inhibition of autophagosome-to-autolysosome fusion or expression of SARS-CoV-2 ORF3a, is sufficient to induce EVP secretion of autophagy cargo receptors. Finally, ATG-dependent EVP secretion buffers against the intracellular accumulation of autophagy cargo receptors when classical autophagic degradation is impaired. Thus, we propose secretory autophagy via EVPs functions as an alternate route to clear sequestered material and maintain proteostasis during endolysosomal dysfunction or impaired autophagosome maturation.

Homonegativity and its relationship to religiosity, nationalism and attachment style.

This study investigated the relationships between negative attitudes towards homosexuals and two traditional ideologies: religiosity and nationalism, and explored the link with attachment style. An Internet survey yielded 290 participants, of highly diverse ages, nationalities, and religious backgrounds. The participants provided demographic details, and completed measures of adult attachment, nationalism, religiosity, and both explicit and implicit measures of homonegativity. The results indicated that both nationalism and religiosity were highly significant predictors of homonegativity. In the religious group, homonegativity and religiosity were positively related. This finding was greater for less securely attached individuals. Avoidance moderated the relationship in religious females, while anxiety moderated the relationship in religious males. No significant attachment moderation was found between the nationalism-homonegativity relationships.

Autophagy suppresses breast cancer metastasis by degrading NBR1.

Macroautophagy/autophagy plays complex, context-dependent roles in cancer. How autophagy governs the emergence of metastatic disease has been incompletely understood. We recently uncovered that genetic autophagy inhibition strongly attenuates primary tumor growth in mammary cancer models, yet paradoxically promotes spontaneous metastasis to the lung and enables the outgrowth of disseminated tumor cells (DTCs) into overt macro-metastases. Furthermore, at both primary and metastatic sites, genetic autophagy inhibition leads to the marked expansion of tumor cells exhibiting aggressive and pro-metastatic basal epithelial differentiation. These pro-metastatic effects of autophagy inhibition are due to the cytosolic accumulation of the autophagy cargo receptor NBR1 in autophagy-deficient tumor cells.

Ribosome profiling reveals a functional role for autophagy in mRNA translational control.

Autophagy promotes protein degradation, and therefore has been proposed to maintain amino acid pools to sustain protein synthesis during metabolic stress. To date, how autophagy influences the protein synthesis landscape in mammalian cells remains unclear. Here, we utilize ribosome profiling to delineate the effects of genetic ablation of the autophagy regulator, ATG12, on translational control. In mammalian cells, genetic loss of autophagy does not impact global rates of cap dependent translation, even under starvation conditions. Instead, autophagy supports the translation of a subset of mRNAs enriched for cell cycle control and DNA damage repair. In particular, we demonstrate that autophagy enables the translation of the DNA damage repair protein BRCA2, which is functionally required to attenuate DNA damage and promote cell survival in response to PARP inhibition. Overall, our findings illuminate that autophagy impacts protein translation and shapes the protein landscape.

Autophagic Degradation of NBR1 Restricts Metastatic Outgrowth during Mammary Tumor Progression.

Although autophagy is being pursued as a therapeutic target in clinical oncology trials, its effects on metastasis, the principal cause of cancer mortality, remain unclear. Here, we utilize mammary cancer models to temporally delete essential autophagy regulators during carcinoma progression. Though genetic ablation of autophagy strongly attenuates primary mammary tumor growth, impaired autophagy promotes spontaneous metastasis and enables the outgrowth of disseminated tumor cells into overt macro-metastases. Transcriptomic analysis reveals that autophagy deficiency elicits a subpopulation of otherwise luminal tumor cells exhibiting basal differentiation traits, which is reversed upon preventing accumulation of the autophagy cargo receptor, Neighbor to BRCA1 (NBR1). Furthermore, pharmacological and genetic induction of autophagy suppresses pro-metastatic differentiation and metastatic outgrowth. Analysis of human breast cancer data reveal that autophagy gene expression inversely correlates with pro-metastatic differentiation signatures and predicts overall and distant metastasis-free survival. Overall, these findings highlight autophagy-dependent control of NBR1 as a key determinant of metastatic progression.

The LC3-conjugation machinery specifies the loading of RNA-binding proteins into extracellular vesicles.

Traditionally viewed as an autodigestive pathway, autophagy also facilitates cellular secretion; however, the mechanisms underlying these processes remain unclear. Here, we demonstrate that components of the autophagy machinery specify secretion within extracellular vesicles (EVs). Using a proximity-dependent biotinylation proteomics strategy, we identify 200 putative targets of LC3-dependent secretion. This secretome consists of a highly interconnected network enriched in RNA-binding proteins (RBPs) and EV cargoes. Proteomic and RNA profiling of EVs identifies diverse RBPs and small non-coding RNAs requiring the LC3-conjugation machinery for packaging and secretion. Focusing on two RBPs, heterogeneous nuclear ribonucleoprotein K (HNRNPK) and scaffold-attachment factor B (SAFB), we demonstrate that these proteins interact with LC3 and are secreted within EVs enriched with lipidated LC3. Furthermore, their secretion requires the LC3-conjugation machinery, neutral sphingomyelinase 2 (nSMase2) and LC3-dependent recruitment of factor associated with nSMase2 activity (FAN). Hence, the LC3-conjugation pathway controls EV cargo loading and secretion.