Read with caution!
This post was written during early stages of trying to understand a complex scientific problem, and we didn't get everything right. The original author no longer endorses the content of this post. It is being left online for historical reasons, but read at your own risk.
“Autophagy and metabolism,” Rabinowitz and White, Science (2010)
Autophagy is a process of self-cannibalization. Cells capture their own cytoplasm and organelles and consume them in lysosomes. The resulting breakdown products are inputs to cellular metabolism, through which they are used to generate energy and to build new proteins and membranes. Autophagy preserves the health of cells and tissues by replacing outdated and damaged cellular components with fresh ones. In starvation, it provides an internal source of nutrients for energy generation and, thus, survival. A powerful promoter of metabolic homeostasis at both the cellular and whole-animal level, autophagy prevents degenerative diseases. It does have a downside, however—cancer cells exploit it to survive in nutrient-poor tumors.
Bulk protein degradation and organelle clearance occurs via two major systems in eukaryotic cells. The ubiquitin-proteasome pathway predominantly degrades short-lived nuclear and cytosolic proteins. The lysosomal-vacuolar pathway degrades larger substrates such as protein complexes and organelles.
Proteasomes are very large protein complexes inside all eukaryotes and archaea, and in some bacteria. In eukaryotes, they are located in the nucleus and the cytoplasm. The main function of the proteasome is to degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks peptide bonds. Enzymes that carry out such reactions are called proteases. Proteasomes are part of a major mechanism by which cells regulate the concentration of particular proteins and degrade misfolded proteins. The degradation process yields peptides of about seven to eight amino acids long, which can then be further degraded into amino acids and used in synthesizing new proteins. Proteins are tagged for degradation with a small protein called ubiquitin. The tagging reaction is catalyzed by enzymes called ubiquitin ligases. Once a protein is tagged with a single ubiquitin molecule, this is a signal to other ligases to attach additional ubiquitin molecules. The result is a polyubiquitin chain that is bound by the proteasome, allowing it to degrade the tagged protein.
In structure, the proteasome is a cylindrical complex containing a “core” of four stacked rings around a central pore. Each ring is composed of seven individual proteins. The inner two rings are made of seven β subunits that contain three to seven protease active sites. These sites are located on the interior surface of the rings, so that the target protein must enter the central pore before it is degraded. The outer two rings each contain seven α subunits whose function is to maintain a “gate” through which proteins enter the barrel. These α subunits are controlled by binding to “cap” structures or regulatory particles that recognize polyubiquitin tags attached to protein substrates and initiate the degradation process. The overall system of ubiquitination and proteasomal degradation is known as the ubiquitin-proteasome system.
“Autophagy induction by trehalose counteracts cellular prion infection,” Aquib et al., Autophagy (2009)
Prion diseases are fatal neurodegenerative and infectious disorders for which no therapeutic or prophylactic regimens exist. In search of cellular mechanisms that play a role in prion diseases and have the potential to interfere with accumulation of intracellular pathological prion protein (PrP(Sc)), we investigated the autophagic pathway and one of its recently published inducers, trehalose. Trehalose, an alpha-linked disaccharide, has been shown to accelerate clearance of mutant huntingtin and alpha-synuclein by activating autophagy, mainly in an mTOR-independent manner. Here, we demonstrate that trehalose can significantly reduce PrP(Sc) in a dose- and time-dependent manner while at the same time it induces autophagy in persistently prion-infected neuronal cells. Inhibition of autophagy, either pharmacologically by known autophagy inhibitors like 3-methyladenine, or genetically by siRNA targeting Atg5, counteracted the anti-prion effect of trehalose. Hence, we provide direct experimental evidence that induction of autophagy mediates enhanced cellular degradation of prions. Similar results were obtained with rapamycin, a known inducer of autophagy, and imatinib, which has been shown to activate autophagosome formation. While induction of autophagy resulted in reduction of PrP(Sc), inhibition of autophagy increased the amounts of cellular PrP(Sc), suggesting that autophagy is involved in the physiological degradation process of cellular PrP(Sc). Preliminary in vivo studies with trehalose in intraperitoneally prion-infected mice did not result in prolongation of incubation times, but demonstrated delayed appearance of PrP(Sc) in the spleen. Overall, our study provides the first experimental evidence for the impact of autophagy in yet another type of neurodegenerative disease, namely prion disease.
“Autophagy, prion infection, and their mutual interactions,” Heiseke et al., Curr. Issues Mol. Biol. (2010)
Prion diseases are infectious and fatal neurodegenerative disorders of man and animals which are characterized by spongiform degeneration in the central nervous system. Prion propagation involves the endocytic pathway and endosomal and lysosomal compartments are implicated in trafficking and re-cycling as well as final degradation of prions. Shifting the equilibrium between propagation and lysosomal clearance to the latter impairs cellular prion load. This and earlier findings of autophagic vacuoles in correlation to prion infections both in in vitro and in vivo studies prompted us and others to analyze the role of autophagy in prion infection. Autophagy is a fundamental cellular bulk degradation process for e.g. organelles or cytoplasmic proteins which has many implications for physiology and patho-physiology of cells and whole organisms. In various neurodegenerative disease models mainly protective functions of autophagy were recently described. In this review, we focus on recent findings which correlate autophagy and its manipulations with prion infection scenarios, and discuss perspectives and future directions. The findings summarized here add to the knowledge of the role of autophagy in neurodegeneration and provide interesting new insight into how non-cytosolic aggregated proteins might be subjected to autophagic clearance.
“Cell death and autophagy in prion diseases (transmissible spongiform encephalopathies), Liberski et al., Folia Neuropathol. (2008)
Neuronal autophagy, like apoptosis, is one of the mechanisms of programmed cell death. In this review, we summarize current information about autophagy in naturally occurring and experimentally induced scrapie, Creutzfeldt-Jakob disease and Gerstmann-Sträussler-Scheinker syndrome against the broad background of neural degenerations in transmissible spongiform encephalopathies (TSEs). Typically a sequence of events is observed: from a part of the neuronal cytoplasm sequestrated by concentric arrays of double membranes (phagophores); through the enclosure of the cytoplasm and membrane proliferation; to a final transformation of the large area of the cytoplasm into a collection of autophagic vacuoles of different sizes. These autophagic vacuoles form not only in neuronal perikarya but also in neurites and synapses. On the basis of ultrastructural studies, we suggest that autophagy may play a major role in transmissible spongiform encephalopathies and may even participate in the formation of spongiform change.
“Prions: New Research,” edited by Brigette v. Doupher (2006) “Autophagy in prion diseases,” Beata Sikorska and Pawel P. Liberski, p 175.
Although the role of autophagy in prion diseases remains unknown, at least three hypotheses must be taken into consideration: 1) autophagy plays a role in removing protein aggregates, 2) autophagy is one route to neuronal death in protein diseases, and 3) it may participate in the formation of spongiform change.
“Autophagy in Immunity and Infection: A novel immune effector,” Deretic (2006), “Autophagy in disease and aging,” Marta Martinez-Vicente et al., p. 69
Although less explored than the neurodegenerative disorders described in the previous sections, several pieces of evidence have connected prion disorders with autophagy. Despite initial reports claiming resistance of PrP to protease cleavage, it has recently been shown that PrP(Sc) can be degraded by lysosomal enzymes, such as cathepsin L and B, thus supporting the participation of the lysosomal system in PrP(Sc) degradation . In addition, accumulation of AVs has been described in neurons from patients and in cells with experimentally induced prion disease [68, 69]. In fact, this intracellular accumulation of AVs, along with other organelle deformities (mitochondrial dilation, enlargement of Golgi and endoplasmic reticulum (ER)) and the absence of typical apoptotic features in the dying neurons, has led to the conclusion that neuronal loss in these disorders occurs via autophagic cell death [68, 70, 71]. Whether this accumulation of AVs occurs only in the advanced stages of the disease, while early activation of autophagy could have a protection effect in the progression of prion diseases, as shown for other neurodegenerative disorders, still needs to be elucidated.
 K.M. Luhr, E.K. Nordstrom, P. Low, K. Kristensson, “Cathepsin B and L are involved in degradation of prions in GT1-1 neuronal cells,” Neuroreport 2004, 1663-1667
 P.P. Liberski, B. Sikorska, J. Bratosiewicz-Wasik, D.C. Gajdusek, P. Brown, “Neuronal cell death in transmissible spongiform encephalopathies (prion diseases) revisited: from apoptosis to autophagy,” Int J Biochem Cell Biol (2004), 36, 2473-2490
 B. Sikorska, P.P. Liberski, P. Giraud, N. Kopp, P. Brown, “Autophagy is a part of ultrastructural synaptic pathology in Creutzfeldt-Jakob disease: a brain biopsy study,” Int J Biochem Cell Biol (2004), 36, 2563-2573
 D. Jesionek-Kupnicka, R. Kordek, J. Buczynski, P.P. Liberski, “Apoptosis in relation to neuronal loss in experimental Creutzfeldt-Jakob disease in mice,” Acta Neurobiol Exp (Wars) 2001, 61, 13-19
 M.B. Graeber, L.B.Moran, “Mechanisms of cell death in neurodegenerative diseases: fashion, fiction, and facts,” Brain Pathol (2002), 12, 385-390
“Bectin 1 controls the levels of p53 by regulating the deubiquitination activity of USP10 and USP13,” Liu et al., Cell (2011)
Autophagy is an important intracellular catabolic mechanism that mediates the degradation of cytoplasmic proteins and organelles. We report a potent small molecule inhibitor of autophagy named “spautin-1” for specific and potent autophagy inhibitor-1. Spautin-1 promotes the degradation of Vps34 PI3 kinase complexes by inhibiting two ubiquitin-specific peptidases, USP10 and USP13, that target the Beclin1 subunit of Vps34 complexes. Beclin1 is a tumor suppressor and frequently monoallelically lost in human cancers. Interestingly, Beclin1 also controls the protein stabilities of USP10 and USP13 by regulating their deubiquitinating activities. Since USP10 mediates the deubiquitination of p53, regulating deubiquitination activity of USP10 and USP13 by Beclin1 provides a mechanism for Beclin1 to control the levels of p53. Our study provides a molecular mechanism involving protein deubiquitination that connects two important tumor suppressors, p53 and Beclin1, and a potent small molecule inhibitor of autophagy as a possible lead compound for developing anticancer drugs.
According to a Wednesday, November 9, 2011 online news article (the story was released by David Cameron of FOCUS, which publishes news from Harvard Medical School, Harvard School of Dental Medicine, and the Harvard School of Public Health):
“Researchers have discovered a small molecule that disables a prized cancer target, one that many pharmaceutical and biotech companies have been investigating for years.
The results were published in the journal “Cell“.
The process, called autophagy, rids cells of debris and is crucial for cell survival.
“Autophagy helps cells survive stress,” said Junying Yuan, Harvard Medical School professor of cell biology and senior author on the paper. “It’s like a recycling process that degrades old proteins into amino acid energy sources enabling cells to survive in difficult circumstances. It’s a turnover mechanism.”
When autophagy falters, life span shortens, and cancer and other diseases, such as neurodegeneration, can ensue. One such defect, in a gene called Beclin1, decreases autophagy in mammalian cells, and researchers have suspected that this leads to increased prostate and breast cancers.
But like so many cancer factors, autophagy can be a double-edged sword.
When a patient is undergoing treatment such as chemotherapy, cancer cells co-opt autophagy and use it to survive the stress of therapy. Researchers have reasoned that in certain clinical settings, briefly disabling autophagy may support and enhance treatment.
For years, pharmaceutical companies have sought to do just that. The challenge lay in identifying the precise target within a protein complex. Yuan and her colleagues developed a cell-based screening platform in which they uncovered a key mechanism of autophagy as well as a small molecule that efficiently blocks the process by degrading the protein complex that autophagy depends on. The protein beclin1, encoded by the Beclin1 gene already linked to autophagy, is a part of this complex.
Drilling deeper, the researchers found that spautin-1 blocked the activity of USP10, a molecule that offers a kind of “stay of execution” for proteins on death row. Proteins marked for disposal are tagged with a marker called ubiquitin, and USP10 often removes this tag from select proteins, sparing them. Removing USP10 leaves these proteins vulnerable.
Beclin1, it turns out, regulates the activity of USP10. And the researchers connected these findings to other studies linking USP10 to p53, a gene widely known to suppress cancer.
“Knocking down Beclin1, which our small molecule does, knocks down USP10, which in turn knocks down p53,” said Yuan. “They are all part of a chain.”
This then explains the earlier observation that mammals with defective Beclin1 experience increased cancer. When beclin1 is diminished, p53, which is downstream, is also diminished, and cancer thrives. However, when Beclin1 is removed altogether, the cell dies. This discovery suggests that selectively targeting autophagy during cancer therapies may greatly benefit patients.
This research was funded by the National Institutes of Health, the Chinese Academy of Sciences, the National Natural Science Foundation of China, and the Harvard University Biomedical Accelerator Fund.
—David Cameron One Response to Long Sought Cancer”