For eukaryotic cells to maintain homeostasis, a balance between protein synthesis and proteolysis must exist. This balance is achieved in a variety of ways. Cells have the ability to control protein synthesis by regulating transcription and translation (1, 2). Similarly, cells can also control protein degradation by using a variety of cellular processes such as lysosomal degradative pathways and the ubiquitin/proteasome pathway (3, 4).
The lysosomal degradative pathways can be separated into endocytosis, macroautophagy, crinophagy, pexophagy, micro-autophagy, and chaperone-mediated autophagy (CMA)1; only the latter pathway does not involve vesicular membrane traffic (5). Autophagy, literally "self-eating," is a cellular process that allows cells to remove proteins, organelles, and foreign bodies from the cytosol and deliver them to lysosomes for degradation.
CMA is a process activated during long term starvation in which cells selectively degrade proteins in order to recycle their amino acids or use them for energy. During nutrient deprivation, substrates that contain a consensus motif related to KFERQ (6) are recognized by a chaperone-cochaperone complex containing the heat shock cognate protein of 70 kDa (hsc70) (7, 8). Once this chaperone-cochaperone complex binds the substrate, it docks on the lysosomal membrane via a receptor known as the lysosomal associated membrane protein 2a (lamp2a) (9). The substrate then is unfolded (10), presumably by the chaperone-cochaperone complex, translocated into the lumen with the help of a lysosomal isoform of hsc70 (lyhsc70) (11), and degraded. Like most organelle protein import pathways, CMA is saturable as well as temperature-dependent (12, 13). The substrates for CMA also compete with one another for binding and import, which provides an experimental method for discovering new substrates (12). There have been several substrates identified for CMA including ribonuclease A (RNase A) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (14, 15). CMA has been reconstituted using lysosomes isolated from human fibroblasts and rat liver, which permits the study of CMA in a more mechanistic fashion (8, 16).
Ketone bodies are produced by the liver during long term starvation in response to rapid lipolysis. Ketone bodies can be utilized by muscle and contribute to the preservation of muscle mass during prolonged survival; they can also be used as an energy source for the brain (17, 18). Ketone bodies are comprised of three biologically active compounds, namely acetoacetate, -hydroxybutyrate (BOH), and acetone (19, 20). Interestingly, the increase in the concentration of circulating ketone bodies parallels the induction of CMA, which is also activated by prolonged starvation (5).
In this paper we demonstrate that treatment of cells with ketone bodies increases the proteolysis of long-lived proteins under conditions in which most proteolysis is due to CMA (28). We also show that the increase in proteolysis observed is at least in part due to the stimulation of CMA. Lastly, we show that ketone bodies induce CMA by oxidizing substrates, permitting them to be recognized by the CMA machinery and imported into the lysosome more efficiently.