Chaperone-mediated autophagy: selectivity pays off

“Three different mechanisms for delivery of autophagic cargo to lysosomes co-exist in most mammalian cells: macroautophagy, microautophagy and chaperone-mediated autophagy (CMA) [6,8] (Box 1).”

“I bet you just pictured a garbage container, didn’t you? This organelle has been associated with cellular cleaning almost since its description more than 50 years ago by Christian de Duve [1].”

“Three different mechanisms for delivery of autophagic cargo to lysosomes co-exist in most mammalian cells: macroautophagy, microautophagy and chaperone-mediated autophagy (CMA) [6,8] (Box 1).”

“The use of blocking antibodies revealed that substrate proteins bind to the lysosomal membrane through the cytosolic tail of LAMP-2A [18].”

“Lastly, CMA is also likely to be altered in Danon’s disease, a type of vacuolar myopathy resulting from mutations in the LAMP2 gene [52].”

“For example, the alterations in macroautophagy and in lysosomal biogenesis observed in Lamp2 null mice [33] and after knocking-out or knocking-down all LAMP-2 isoforms in cultured cells [34,35], cannot be reproduced by blocking only LAMP-2A in cultured cells [25].”

“Alphasynuclein can be degraded by the ubiquitin proteasome system, CMA and macroautophagy [43,44].”

“It is likely that LAMP-2A participates in other cellular functions common to all LAMP-2 protein variants with which LAMP-2A shares a common luminal region [61].”

“those containing HSC70 in their lumen, through still unknown mechanisms [37].”

“Instead, pathogenic alpha-synucleins remain tightly bound to the CMA receptor, preventing the translocation of other CMA substrates into lysosomes [43,45].”

“A mutant cleaved form of Tau, the protein that organizes into toxic fibrillar structures in Alzheimer’s disease (AD), is also delivered to lysosomes via CMA, and similar to the abnormal alpha-synucleins, mutant Tau fails to translocate properly [47].”

“For example, some motifs located in the core of the protein will only be exposed upon unfolding [28]; other motifs placed in regions of protein/protein interaction will become accessible when the protein is no longer part of a complex [60]; lastly, in some enzymes the motifs are close to their catalytic site and only…”

“For most of these functions, HSC70 binds to stretches of hydrophobic residues in the substrates; however, to date, the KFERQ-like motif is the only region recognized by HSC70 in CMA substrates [12].”

“For example, RNA interference against LAMP-2A, which blocks CMA results in constitutive activation of macroautophagy and vice versa; however, both pathways are non-redundant, and the lack of the defective pathway becomes evident in response to an additional stressor [25,27].”

“Similarly, the age-dependent decline in CMA activity could be an aggravating factor in most of these neurodegenerative disorders [39].”

“We developed a bi-transgenic mouse model expressing an additional copy of LAMP-2A in liver that can be activated in middle life to supplement the reduced levels of endogenous protein [38].”

“The first function proposed for CMA was to provide an alternative source of amino acids under conditions of prolonged starvation [9,10,24].”

“For example, RNA interference against LAMP-2A, which blocks CMA results in constitutive activation of macroautophagy and vice versa; however, both pathways are non-redundant, and the lack of the defective pathway becomes evident in response to an additional stressor [25,27].”

“HSC70 is a multifunctional chaperone that also contributes to the disassembly of clathrin from endocytic pits, folding of newly synthesized proteins, refolding of unfolded proteins and, as a partner of HSP90, to the stabilization of proteins in transient unstable conformations [14].”

“Some recent studies propose that the carboxyl terminus of HSP70-interacting protein (CHIP), another cytosolic chaperone that contributes to the degradation of cytosolic proteins by the proteasome, might also target proteins for lysosomal degradation [15].”

“Blocking antibodies against luminal HSC70 prevented substrate translocation, supporting that this chaperone might act by “pulling” the substrates inside the lysosomal lumen as they cross the membrane [22].”

“In professional antigen presenting cells, CMA participates in the presentation of cytoplasmic peptides by class II molecules contributing to organism autoimmunity [30].”

“Recently, malfunctioning of CMA has been connected to a growing number of human disorders [42].”

“Furthermore, not only pathogenic forms of alphasynuclein, but even high levels (overexpression) of the wild type form of this protein has recently been found to hinder the degradation of neuronal proteins by CMA [31].”

“Formation of the limiting membrane also requires the interaction between a protein and a specific lipid molecule (phosphatidyl ethanolamine), also regulated by conjugating enzymes [7].”

“Luminal HSC70 is necessary for CMA, and in fact, the subset of lysosomes able to perform CMA is often identified in cells by the presence of HSC70 in their lumen [23].”

“Conditions associated with enhanced CMA always require high LAMP-2A levels, which can be attained by transcriptional upregulation of the Lamp2 gene [28], but more often through changes in the degradation and lysosomal distribution of LAMP-2A [19].”

“Numerous reviews have summarized the recent findings on the coordinated orchestration of macroautophagy, the mechanisms that regulate this process, and its important and growing number of physiological functions [5-8].”

“For example, the alterations in macroautophagy and in lysosomal biogenesis observed in Lamp2 null mice [33] and after knocking-out or knocking-down all LAMP-2 isoforms in cultured cells [34,35], cannot be reproduced by blocking only LAMP-2A in cultured cells [25].”

“Tracking of individual proteins undergoing selective lysosomal degradation and analyzing their sequences led to the identification of both a targeting motif in all of the substrates for this pathway and a chaperone able to recognize this motif [11,12].”

“Chimeras of CMA substrates with a protein unable to unfold can bind to the lysosomal membrane but do not translocate, suggesting that unfolding occurs after binding but before translocation [17].”

“The connections between CMA and PD have further expanded with the discovery that mutant forms of the ubiquitin carboxyl-terminal esterase L1 (UCHL-1), another pathogenic protein in PD, interact abnormally with LAMP-2A leading to CMA blockage [46].”

“Enhanced recruitment of LAMP-2A to these areas of degradation at the lysosomal membrane reduces levels of this receptor and downregulates CMA [21].”

“Instead, the stability of the CMA receptor is markedly compromised once it reaches the lysosome in old organisms because a large percentage of lysosomal LAMP-2A remains in the lysosomal lumen where it undergoes massive unregulated degradation [40].”

“In contrast to macro- and microautophagy, delivery of cargo via CMA does not require formation of intermediate vesicle compartments, membrane fusion or membrane deformity of any kind [57,58].”

“Degradation is initiated through dual cleavage by a pair of lysosomal proteases, cathepsin A and a yet unidentified metalloprotease, followed by the release of the truncated form of LAMP-2A in the lumen and its complete degradation by resident hydrolases [50].”

“Macroautophagy is activated in most cells during the first hours of starvation, reaches its maximum activity at about 6 hours and then gradually declines [25,26].”

“A recent study reported abnormal upregulation of macroautophagy in mice with accelerated aging that might contribute to their systemic degeneration [41].”

“Interestingly, increased levels of oxidized proteins might also contribute to keeping CMA upregulated during prolonged starvation, because the increased levels of circulating ketone bodies in these conditions promotes protein oxidation [29].”

“For example, some motifs located in the core of the protein will only be exposed upon unfolding [28]; other motifs placed in regions of protein/protein interaction will become accessible when the protein is no longer part of a complex [60]; lastly, in some enzymes the motifs are close to their catalytic site and only…”

“The observed changes in CMA activity in the affected cells could be secondary to the changes in pH; however, because their lysosomes contain lower amounts of cytosolic chaperones, it is also possible that TRPML1 might participate in lysosomal docking of CMA chaperones [51].”

“Early studies in cultured primary fibroblasts revealed that the pool of cytosolic proteins whose lysosomal degradation was accelerated upon serum removal was degraded at slower rates as cells become senescent [36].”

“protein also implicated in the pathogenesis of this disease, undergoes CMA [48].”

“Decreased CMA activity under these conditions leads to high levels of the nuclear transcription factor Pax2, essential in regulating cellular growth in the kidney [54].”

“Modulation of kidney growth is a specialized function attributed to CMA through the degradation of a specific protein, the transcription factor Pax2, in renal tubular cells [32].”

“Most of the early studies on autophagy were performed in rat liver, tracking the degradation of the soluble pool of cytosolic protein or measuring changes in the size and volume of the lysosomal system [2-4].”

“To date, only one membrane protein, the epidermal growth factor receptor, has been proposed to undergo lysosomal degradation in a manner dependent on the receptor for CMA [59]; however, the requirement for protein unfolding, essential to define this process as CMA, has not yet been determined for the epidermal growth…”

“The first function proposed for CMA was to provide an alternative source of amino acids under conditions of prolonged starvation [9,10,24].”

“For example, the alterations in macroautophagy and in lysosomal biogenesis observed in Lamp2 null mice [33] and after knocking-out or knocking-down all LAMP-2 isoforms in cultured cells [34,35], cannot be reproduced by blocking only LAMP-2A in cultured cells [25].”

“Lysosomal storage disorders (LSD) are a group of genetic diseases in which the lack of enzymatic activity in lysosomes leads to massive accumulation of the substrate of that enzyme, resulting in lysosomal engorgement [49].”

“To date, three autophagy-related pathways have been described in higher eukaryotes: macroautophagy (MA), chaperone-mediated autophagy (CMA), andmicroautophagy (Cuervo, 2010; Mizushima et al., 2008).”

“In the third common type of autophagy, CMA, cargo is not sequestered but is instead selectively recognized by a complex of cytosolic chaperones that mediates its delivery to a receptor at the lysosomal membran”