Daniel J. Klionsky

Autophagy fights disease through cellular self-digestion

Autophagy, or cellular self-digestion, is a cellular pathway involved in protein and organelle degradation, with an astonishing number of connections to human disease and physiology. For example, autophagic dysfunction is associated with cancer, neurodegeneration, microbial infection and ageing. Paradoxically, although autophagy is primarily a protective process for the cell, it can also play a role in cell death. Understanding autophagy may ultimately allow scientists and clinicians to harness this process for the purpose of improving human health.

Development by self-digestion: Molecular mechanisms and biological functions of autophagy

Autophagy is the major cellular pathway for the degradation of long-lived proteins and cytoplasmic organelles. It involves the rearrangement of subcellular membranes to sequester cargo for delivery to the lysosome where the sequestered material is degraded and recycled. For many decades, it has been known that autophagy occurs in a wide range of eukaryotic organisms and in multiple different cell types during starvation, cellular and tissue remodeling, and cell death. However, until recently, the functions of autophagy in normal development were largely unknown. The identification of a set of evolutionarily conserved genes that are essential for autophagy has opened up new frontiers for deciphering the role of autophagy in diverse biological processes. In this review, we summarize our current knowledge about the molecular machinery of autophagy and the role of the autophagic machinery in eukaryotic development.

Regulation Mechanisms and Signaling Pathways of Autophagy

Autophagy is a process of self-degradation of cellular components in which double-membrane autophagosomes sequester organelles or portions of cytosol and fuse with lysosomes or vacuoles for breakdown by resident hydrolases. Autophagy is upregulated in response to extra- or intracellular stress and signals such as starvation, growth factor deprivation, ER stress, and pathogen infection. Defective autophagy plays a significant role in human pathologies, including cancer, neurodegeneration, and infectious diseases. We present our current knowledge on the key genes composing the autophagy machinery in eukaryotes from yeast to mammalian cells and the signaling pathways that sense the status of different types of stress and induce autophagy for cell survival and homeostasis. We also review the recent advances on the molecular mechanisms that regulate the autophagy machinery at various levels, from transcriptional activation to post-translational protein modification.

Autophagosome formation: Core machinery and adaptations

Eukaryotic cells employ autophagy to degrade damaged or obsolete organelles and proteins. Central to this process is the formation of autophagosomes, double-membrane vesicles responsible for delivering cytoplasmic material to lysosomes. In the past decade many autophagy-related genes, ATG, have been identified that are required for selective and/or nonselective autophagic functions. In all types of autophagy, a core molecular machinery has a critical role in forming sequestering vesicles, the autophagosome, which is the hallmark morphological feature of this dynamic process. Additional components allow autophagy to adapt to the changing needs of the cell.

Mammalian autophagy: core molecular machinery and signaling regulation

Autophagy, a cellular catabolic pathway, is evolutionarily conserved from yeast to mammals. Central to this process is the formation of autophagosomes, double-membrane vesicles responsible for delivering long-lived proteins and excess or damaged organelle into the lysosome for degradation and reuse of the resulting macromolecules. In addition to the hallmark discovery of core molecular machinery components involved in autophagosome formation, complex signaling cascades controlling autophagy have also begun to emerge, with mTOR as a central but far from exclusive player. Malfunction of autophagy has been linked to a wide range of human pathologies, including cancer, neurodegeneration, and pathogen infection. Here we highlight the recent advances in identifying and understanding the core molecular machinery and signaling pathways that are involved in mammalian autophagy.

A protein conjugation system essential for autophagy

Autophagy is a process for the bulk degradation of proteins, in which cytoplasmic components of the cell are enclosed by double-membrane structures known as autophagosomes for delivery to lysosomes or vacuoles for degradation. This process is crucial for survival during starvation and cell differentiation. No molecules have been identified that are involved in autophagy in higher eukaryotes. We have isolated 14 autophagy-defective (apg) mutants of the yeast Saccharomyces cerevisiae and examined the autophagic process at the molecular level. We show here that a unique covalent-modification system is essential for autophagy to occur. The carboxy-terminal glycine residue of Apg12, a 186-amino-acid protein, is conjugated to a lysine at residue 149 of Apg5, a 294-amino-acid protein. Of the apg mutants, we found that apg7 and apg10 were unable to form an Apg5/Apg12 conjugate. By cloning APG7, we discovered that Apg7 is a ubiquitin-E1-like enzyme. This conjugation can be reconstituted in vitro and depends on ATP. To our knowledge, this is the first report of a protein unrelated to ubiquitin that uses a ubiquitination-like conjugation system. Furthermore, Apg5 and Apg12 have mammalian homologues, suggesting that this new modification system is conserved from yeast to mammalian cells.

Autophagy: molecular machinery for self-eating

Autophagy is a highly conserved process in eukaryotes in which the cytoplasm, including excess or aberrant organelles, is sequestered into double-membrane vesicles and delivered to the degradative organelle, the lysosome/vacuole, for breakdown and eventual recycling of the resulting macromolecules. This process has an important role in various biological events such as adaptation to changing environmental conditions, cellular remodeling during development and differentiation, and determination of lifespan. Auto-phagy is also involved in preventing certain types of disease, although it may contribute to some pathologies. Recent studies have identified many components that are required to drive this complicated cellular process. Auto-phagy-related genes were first identified in yeast, but homologs are found in all eukaryotes. Analyses in a range of model systems have provided huge advances toward understanding the molecular basis of autophagy. Here we review our current knowledge on the machinery and molecular mechanism of autophagy.

A unified nomenclature for yeast autophagy-related genes

The authors would like to thank Drs. Jan A.K.W. Kiel, Ida J. van der Klei, Beth Levine, Fulvio Reggiori, and Takahiro Shintani for helpful comments on the manuscript, and the many researchers in the yeast field who have agreed to changes in the standard names of various genes.

Potential therapeutic applications of autophagy

Autophagy is a dynamic process of subcellular degradation, which has recently sparked great interest as it is now recognized to be involved in various developmental processes and various diseases including cancer and neurodegeneration. Autophagy can function as a cytoprotective mechanism; however, it also has the capacity to cause cell death. A better understanding of autophagy is needed to allow its manipulation for therapeutic purposes, and new insights into the molecular mechanisms of autophagy are now leading to the discovery of exciting new potential drug targets.

The molecular machinery of autophagy: unanswered questions

Autophagy is a process in which cytosol and organelles are sequestered within double-membrane vesicles that deliver the contents to the lysosome/vacuole for degradation and recycling of the resulting macromolecules. It plays an important role in the cellular response to stress, is involved in various developmental pathways and functions in tumor suppression, resistance to pathogens and extension of lifespan. Conversely, autophagy may be associated with certain myopathies and neurodegenerative conditions. Substantial progress has been made in identifying the proteins required for autophagy and in understanding its molecular basis; however, many questions remain. For example, Tor is one of the key regulatory proteins at the induction step that controls the function of a complex including Atg1 kinase, but the target of Atg1 is not known. Although autophagy is generally considered to be nonspecific, there are specific types of autophagy that utilize receptor and adaptor proteins such as Atg11; however, the means by which Atg11 connects the cargo with the sequestering vesicle, the autophagosome, is not understood. Formation of the autophagosome is a complex process and neither the mechanism of vesicle formation nor the donor membrane origin is known. The final breakdown of the sequestered cargo relies on well-characterized lysosomal/vacuolar proteases; the roles of lipases, by contrast, have not been elucidated, and we do not know how the integrity of the lysosome/vacuole membrane is maintained during degradation.