What Is Autophagy?

Autophagy is the natural, regulated mechanism of the cell that disassembles unnecessary or dysfunctional components.

It allows the orderly degradation and recycling of cellular components. In macroautophagy, targeted cytoplasmic constituents are isolated from the rest of the cell within a double-membraned vesicle known as an autophagosome. The autophagosome eventually fuses with lysosomes and the contents are degraded and recycled. Three forms are commonly described: macroautophagy, microautophagy, and chaperone-mediated autophagy (CMA).

In disease, it has been seen as an adaptive response to stress, which promotes survival, whereas in other cases it appears to promote cell death and morbidity. In the extreme case of starvation, the breakdown of cellular components promotes cellular survival by maintaining cellular energy levels.

The term was coined by Belgian biochemist Christian de Duve in 1963. The identification of autophagy-related genes in yeast in the 1990s let researchers figure out the mechanisms of autophagy and led to the award of the 2016 Nobel Prize in Physiology or Medicine to Japanese autophagy researcher Yoshinori Ohsumi.

Process And Pathways

There are three main types (macroautophagy, microautophagy and Chaperone mediated autophagy), these are mediated by the autophagy-related genes and their associated enzymes.

Macroautophagy is then divided into bulk and selective types. The selective form is involved in oragenelles; mitophagy, lipophagy, pexophagy, chlorophagy, ribophagy and others.

Macroautophagy is the main pathway, used primarily to eradicate damaged cell organelles or unused proteins. First the phagophore engulfs the material that needs to be degraded, which forms a double membrane known as an autophagosome, around the organelle marked for destruction. The autophagosome then travels through the cytoplasm of the cell to a lysosome, and the two organelles fuse. Within the lysosome, the contents of the autophagosome are degraded via acidic lysosomal hydrolase.

Microautophagy, on the other hand, involves the direct engulfment of cytoplasmic material into the lysosome. This occurs by invagination, meaning the inward folding of the lysosomal membrane, or cellular protrusion.

Chaperone-mediated autophagy, or CMA, is a very complex and specific pathway, which involves the recognition by the hsc70-containing complex. This means that a protein must contain the recognition site for this hsc70 complex which will allow it to bind to this chaperone, forming the CMA- substrate/chaperone complex. This complex then moves to the lysosomal membrane-bound protein that will recognise and bind with the CMA receptor, allowing it to enter the cell.

Upon recognition, the substrate protein gets unfolded and it is translocated across the lysosome membrane with the assistance of the lysosomal hsc70 chaperone. CMA is significantly different from other types because it translocates protein material in a one by one manner, and it is extremely selective about what material crosses the lysosomal barrier.

Mitophagy is the selective degradation of mitochondria by autophagy. It often occurs to defective mitochondria following damage or stress. Mitophagy promotes turnover of mitochondria and prevents accumulation of dysfunctional mitochondria which can lead to cellular degeneration. It is mediated by Atg32 (in yeast) and NIX and its regulator BNIP3 in mammals. Mitophagy is regulated by PINK1 and parkin proteins. The occurrence of mitophagy is not limited to the damaged mitochondria but also involves undamaged ones.

Lipophagy is the degradation of lipids by autophagy, a function which has been shown to exist in both animal and fungal cells. The role of lipophagy in plant cells however remains elusive. In lipophagy the target are lipid structures called lipid droplets (LDs), spheric “organelles” with a core of mainly triacylglycerols (TAGs) and a unilayer of phospholipids and membrane proteins. In animal cells the main lipophagic pathway is via the engulfment of LDs by the phagophore, macroautophagy. In fungal cells on the other hand microplipophagy constitutes the main pathway and is especially well studied in yeast Saccharomyces cerevisiae.

Functions Of Autophagy

Nutrient Starvation

Autophagy has roles in various cellular functions. One particular example is in yeasts, where the nutrient starvation induces a high level of the process. This allows unneeded proteins to be degraded and the amino acids recycled for the synthesis of proteins that are essential for survival.

In higher eukaryotes, the process is induced in response to the nutrient depletion that occurs in animals at birth after severing off the trans-placental food supply, as well as that of nutrient starved cultured cells and tissues. Mutant yeast cells that have a reduced autophagic capability rapidly perish in nutrition-deficient conditions.

Studies on the apg mutants suggest that autophagy via autophagic bodies is indispensable for protein degradation in the vacuoles under starvation conditions, and that at least 15 APG genes are involved in autophagy in yeast. A gene known as ATG7 has been implicated in nutrient-mediated autophagy, as mice studies have shown that starvation-induced autophagy was impaired in atg7-deficient mice.

Xenophagy

In microbiology, xenophagy is the autophagic degradation of infectious particles. Cellular autophagic machinery also play an important role in innate immunity.

Intracellular pathogens, such as Mycobacterium tuberculosis (the bacterium which is responsible for tuberculosis) are targeted for degradation by the same cellular machinery and regulatory mechanisms that target host mitochondria for degradation. Incidentally, this is further evidence for the endosymbiotic hypothesis.

This process generally leads to the destruction of the invasive organism, although some bacteria can block the maturation of phagosomes into degradative organelles called phagolysosomes. Stimulation of autophagy in infected cells can help overcome this phenomenon, restoring pathogen degradation.

Infection

Vesicular stomatitis virus is believed to be taken up by the autophagosome from the cytosol and translocated to the endosomes where detection takes place by a member of the PRRs called toll-like receptor 7, detecting single stranded RNA. Following activation of the toll-like receptor, intracellular signaling cascades are initiated, leading to induction of interferon and other antiviral cytokines.

A subset of viruses and bacteria subvert the autophagic pathway to promote their own replication. Galectin-8 has recently been identified as an intracellular “danger receptor”, able to initiate autophagy against intracellular pathogens. When galectin-8 binds to a damaged vacuole, it recruits autophagy adaptor such as NDP52 leading to the formation of an autophagosome and bacterial degradation.

Repair Mechanism

Autophagy degrades damaged organelles, cell membranes and proteins, and the failure of autophagy is thought to be one of the main reasons for the accumulation of damaged cells and aging.

Programmed Cell Death

One of the mechanisms of programmed cell death (PCD) is associated with the appearance of autophagosomes and depends on autophagous proteins. This form of cell death most likely corresponds to a process that has been morphologically defined as autophagic PCD. One question that constantly arises, however, is whether autophagic activity in dying cells is the cause of death or is actually an attempt to prevent it.

Morphological and histochemical studies so far did not prove a causative relationship between the autophagic process and cell death. In fact, there have recently been strong arguments that autophagic activity in dying cells might actually be a survival mechanism.

Studies of the metamorphosis of insects have shown cells undergoing a form of PCD that appears distinct from other forms; these have been proposed as examples of autophagic cell death.

Recent pharmacological and biochemical studies have proposed that survival and lethal autophagy can be distinguished by the type and degree of regulatory signaling during stress particularly after viral infection. Although promising, these findings have not been examined in non-viral systems.

Cancer

Oftentimes, cancer occurs when several different pathways that regulate cell differentiation are disturbed. Autophagy plays an important role in cancer – both in protecting against cancer as well as potentially contributing to the growth of cancer.

The process can contribute to cancer by promoting survival of tumor cells that have been starved, or that degrade apoptotic mediators through autophagy: in such cases, use of inhibitors of the late stages of autophagy (such as chloroquine), on the cells that use autophagy to survive, increases the number of cancer cells killed by antineoplastic drugs.

The role of autophagy in cancer is one that has been highly researched and reviewed. There is evidence that emphasizes the role of autophagy both as a tumor suppressor as well as a factor in tumor cell survival. However, recent research has been able to show that the process is more likely to be used as a tumor suppressor according to several models.

Tumor Suppressor

Several experiments have been done with mice and varying Beclin1, a protein that regulates autophagy.

When the Beclin1 gene was altered to be heterozygous (Beclin 1+/-), the mice were found to be tumor prone. However, when Beclin1 was overexpressed, tumor development was inhibited.

Care should be exercised when interpreting phenotypes of beclin mutants and attributing the observations to a defect in autophagy, however: Beclin1 is generally required for phosphatidylinositol 3- phosphate production and as such it affects numerous lysosomal and endosomal functions, including endocytosis and endocytic degradation of activated growth factor receptors.

In support of the possibility that Beclin1 affects cancer development through an autophagy-independent pathway is the fact that core autophagy factors which are not known to affect other cellular processes and are definitely not known to affect cell proliferation and cell death, such as Atg7 or Atg5, show a much different phenotype when the respective gene is knocked out, which does not include tumor formation. In addition, full knockout of Beclin1 is embryonic lethal whereas knockout of Atg7 or Atg5 is not.

Necrosis and chronic inflammation also has been shown to be limited through autophagy which helps protect against the formation of tumor cells. Thus these experiments show autophagy’s role as a tumor suppressor.

Tumor Cell Survival

Alternatively, autophagy has also been shown to play a large role in tumor cell survival. In cancerous cells, the process is used as a way to deal with stress on the cell. Once these autophagy related genes were inhibited, cell death was potentiated. The increase in metabolic energy is offset by autophagy functions. These metabolic stresses include hypoxia, nutrient deprivation, and an increase in proliferation.

These stresses activate autophagy in order to recycle ATP and maintain survival of the cancerous cells. Autophagy has been shown to enable continued growth of tumor cells by maintaining cellular energy production. By inhibiting autophagy genes in these tumors cells, regression of the tumor and extended survival of the organs affected by the tumors were found.

Furthermore, inhibition of autophagy has also been shown to enhance the effectiveness of anticancer therapies.

Therapeutic Target

New developments in research have found that targeted autophagy may be a viable therapeutic solution in fighting cancer. As discussed above, the process plays both a role in tumor suppression and tumor cell survival.

Thus, the qualities of autophagy can be used as a strategy for cancer prevention. The first strategy is to induce it and enhance its tumor suppression attributes. The second strategy is to inhibit the process and thus induce apoptosis.

The first strategy has been tested by looking at dose-response anti-tumor effects during autophagy-induced therapies. These therapies have shown that autophagy increases in a dose-dependent manner. This is directly related to the growth of cancer cells in a dose-dependent manner as well. This data supports the development of therapies that will encourage autophagy. Secondly, inhibiting the protein pathways directly known to induce the process may also serve as an anticancer therapy.

The second strategy is based on the idea that it is a protein degradation system used to maintain homeostasis and the findings that inhibition of autophagy often leads to apoptosis. Inhibition is riskier as it may lead to cell survival instead of the desired cell death.

Roberta A. Gottlieb (Editor)
Autophagy in Health and Disease
Academic Press ISBN: 978-0123851017