Autophagy independent roles of the core ATG proteins

By Christina Towers, PhD.

Autophagy and ATG Proteins

Autophagy is a nutrient recycling process that cells use to fuel metabolism, particularly in response to nutrient deprivation.  It is critical for removal of damaged proteins as well as clearing certain infections.  It is a complex process that involves over 20 core autophagy genes (ATGs), most of which were originally identified in yeast but the mammalian homologs have now been well characterized.

The formation of a double membrane vesicle termed the phagophore is initiated by the ULK complex where the protein kinases ULK1 and ULK2 interact with FIP200 to subsequently activate the BECN complex. This involves interactions between BECN1, BH3-containing proteins and the PI3-kinase (VPS34).  Phagophore elongation is dependent on two ubiquitin-like conjugation systems where the E1 enzyme ATG7 conjugates ATG5 to ATG12 and the ATG5-12 conjugate acts as an E3 enzyme to conjugate the GABARAP protein, LC3, to phosphoethanolamine (PE). LC3-PE is not only necessary but is itself incorporated into the elongating double membrane structure deemed an autophagosome once closure has been completed.

Immunocytochemistry/Immunofluorescence: Beclin 1 Antibody [NB500-249] - Beclin 1/ATG6 Antibody [NB500-249] - Beclin 1 antibody was tested in HeLa cells with Dylight 488 (green). Nuclei and alpha-tubulin were counterstained with DAPI (blue) and Dylight 550 (red).

Snare proteins such as STX17 facilitate autophagosome-lysosome fusion, where lysosomal proteins like LAMP2 are also critical.  Once fusion is complete, pH dependent proteases can then degrade the engulfed cytoplasmic material¹.  The majority of in-vitro and in-vivo studies evaluating the role of autophagy in the pathology of human related diseases such as cancer and neurological diseases have focused on genetic ablation of one or two ATGs, most notably ATG5 or ATG7.  However, given the complexity of the process, one criticism of the field could be why not target the other genes of the pathway? This critique is then magnified with a large body of literature now indicating autophagy-independent functions for almost every core ATG in the pathway.

Non-Autophagy Roles of ATG Proteins

1. ATG7 has been implicated in nutrient deprivation induced cell cycle arrest via direct interaction and induction of p53. Most interestingly, this role is independent from its function in autophagy as its E1 enzymatic activity is not necessary for the p53 interaction².

2. BECN1, which was originally identified as the first link between autophagy and tumor suppression, has now been linked to chromosomal instability due to autophagy independent interactions with kinetochore assembly³. 

3. FIP200, which is ablated in mouse models via point mutations that eliminate its autophagic role, can also play a critical role in TNF alpha-induced apoptosis⁴.

4. ATG5 and ATG12 have also been implicated in mitochondrial regulated apoptosis and interaction with BCL2 protein family members⁵.

Additionally, there are even more autophagy independent, yet still autophagy-related mechanisms that involve parts of the pathway. For example, LC3-associated phagocytosis (LAP) is dependent on most of the core ATG proteins, however unlike canonical autophagy LAP is also dependent upon an additional component of the BECN1 complex, RUBICON⁶.

Together these studies highlight the need to further understand the autophagy independent and/or related functions of the core autophagic machinery.  This is especially important for the interpretation of current and on-going studies that are being used as rational to target autophagy (either inhibition or induction) for treatment in patients. 

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Christina Towers, PhD
University of Colorado (AMC)
Dr. Towers studies the roles of autophagy, apoptosis and cell death in cancer.

References

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3. Fremont, S. et al. Beclin-1 is required for chromosome congression and proper outer kinetochore assembly. EMBO Rep 14, 364-372, doi:10.1038/embor.2013.23 (2013).
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6. Martinez, J. et al. Molecular characterization of LC3-associated phagocytosis reveals distinct roles for Rubicon, NOX2 and autophagy proteins. Nature cell biology 17, 893-906, doi:10.1038/ncb3192 (2015).