The different autophagy degradation pathways and neurodegeneration

Abstract

The term autophagy encompasses different pathways that route cytoplasmic material to lysosomes for degradation and includes macroautophagy, chaperone-mediated autophagy, and microautophagy. Since these pathways are crucial for degradation of aggregate-prone proteins and dysfunctional organelles such as mitochondria, they help to maintain cellular homeostasis. As post-mitotic neurons cannot dilute unwanted protein and organelle accumulation by cell division, the nervous system is particularly dependent on autophagic pathways. This dependence may be a vulnerability as people age and these processes become less effective in the brain. Here, we will review how the different autophagic pathways may protect against neurodegeneration, giving examples of both polygenic and monogenic diseases. We have considered how autophagy may have roles in normal CNS functions and the relationships between these degradative pathways and different types of programmed cell death. Finally, we will provide an overview of recently described strategies for upregulating autophagic pathways for therapeutic purposes.

Figure 1.. Schematic representation of macroautophagy:

Cell components to be degraded are engulfed in a double membraned structure called the phagophore, the edges of which elongate and close to form autophagosomes. These ultimately fuse with the lysosomal membrane for cargo degradation. A. Early steps in macroautophagy involve 2 ubiquitin-like conjugation cascades. Conjugation I leads to the formation of the Atg5-Atg12 conjugate mediated by ATG7 (E1-like) and ATG10 (E2-like). This then forms a complex with ATG16L1. During Conjugation II, ATG4 cleaves the C-terminus of LC3 generating LC3-I whose C-terminal glycine can be conjugated to phosphatidylethanolamine (PE) by ATG7 (E1-like), ATG3 (E2-like) and ATG5-ATG12/ATG16L1 (E3-like) (generating lipidated LC3 (LC3-II)). The other ATG8 family members (GABARAPs) use the same machinery to enable their conjugation to PE as LC3 proteins. The sites of LC3 conjugation to membranes are determined by the ATG5-ATG12-ATG16L1 complex, which localizes to the surface of the forming phagophore by interacting with WIPI2. WIPI2 is recruited to these membranes by binding both to phosphatidylinositol 3-phosphate (PI3P) and RAB11A. B. During autophagosome formation, the phagophore double membrane elongates and fuses to form a double-membraned vesicle termed the autophagosome. Cargo within autophagosomes can be trapped in a (i) bulk or (ii) selective manner by autophagy cargo receptors, such as P62, leading to the selective autophagy of specific substrates. Completion of vesicle closure to engulf regions of cytoplasm and organelles or to engulf specific cargoes such as aggregates (aggrephagy), mitochondria (mitophagy) or ribosomes (ribophagy) is followed by release from the recycling endosome-RAB11A platform to which the LC3 conjugates. Finally, the autophagosome outer membrane fuses with the lysosomal membrane and cargo is released for complete degradation in the lysosomal lumen.

Figure 2.. Schematic representation of chaperone-mediated autophagy (CMA) and endosomal microautophagy (eMI).

The first steps for cargo recognition are shared by CMA and eMI and are mediated by the binding of HSC70 to a targeting motif in the protein sequence biochemically related to the pentapeptide KFERQ. The inset (top right) highlights the chemical requirements for KFERQ motifs and the post-translational modifications such as phosphorylation or acetylation that can generate motifs by providing the missing charges. This motif is necessary and sufficient for CMA, whereas it is necessary but insufficient for eMI. Additional, as yet unknown, mediators are required for eMI targeting. HSC70 binds to the surface of late endosomes via phosphoserine and triggers assembly of the ESCRT machinery for internalization of substrate into intraluminal vesicles. In CMA, binding of the HSC70/substrate complex to LAMP2A at the lysosomal membrane triggers its multimerization to form a translocation complex that mediates the internalization of the substrate protein into the lumen for degradation. LAMP2A is actively disassembled from the complex to initiate a new cycle of binding/internalization. LAMP2A also mobilizes laterally to incorporate into lipid microdomains for its own degradation triggered by cathepsin A (CTSA). Phosph. gen.: phosphorylation generated. Acetyl. gen: Acetylation generated.

Figure 3.. Schematic representation of microautophagy:

Bulk of proteins and cells components such as organelles (1. In bulk) can be integrated into lysosomes and late endosomes directly through invaginations at the lysosomal membrane. Cytosolic proteins targeted by Hsc70 can be also selectively degraded by its internalization into late endosome invaginations in a process known as Endosomal microautophagy (2).

Figure 4.. Overview of the role of macroautophagy in the nervous system in health and neurodegeneration.

Autophagy is fundamental to sustain the homoeostasis and function of the CNS. Perturbations in the macroautophagy pathway at different stages have been observed during neurodegeneration and distinct disease-associated genes are also key contributors to macroautophagy dysfunction. Defective autophagy compromises protein clearance and organelle turnover, leading to the accumulation of toxic proteins and damaged cellular components that finally alter neuronal function and induce neuronal loss.

Figure 5.. Autophagy tethering compounds (ATTEC) and the autophagy-targeting chimera (AUTAC) system.

A) ATTEC molecules tether the protein of interest to the autophagosomes by direct binding to the protein of interest and to LC3. A proof-of-concept study using the mutant HTT protein (mHTT) demonstrated that these compounds can degrade mHTT both in cells and in vivo in animal models and demonstrated targeting of mHTT to autophagosomes for subsequent degradation without influencing autophagy activity per se. B) AUTAC technology has a similar design to the PROTAC technology and both use ubiquitination to target proteins for degradation. The AUTAC molecule contains a degradation tag (a guanine derivative called FBnG) which induces K63 polyubiquitination and a ligand which binds to the target protein to provide target specificity. The resulting K63 ubiquitination targets the labelled protein for degradation via macroautophagy.