Role of CALCOCO1 in scaling down endoplasmic reticulum and Golgi by autophagy

Abstract

Endoplasmic reticulum (ER) and Golgi apparatus are key organelles in the synthesis, modification and trafficking of proteins in eukaryotic cells. In response to stress stimuli such as nutrients deprivation, accumulation of misfolded proteins or exposure to chemicals, the ER increases in size through increased synthesis of its components to counteract the stress. Similarly, Golgi response to stress increases synthesis of its components to augment its functions. The excess ER components are scaled down by ER-phagy to restore the physiological size. ER-phagy is a form of autophagy that targets specific portions of the ER for degradation in the lysosome. The degradation is mediated by adaptor molecules called ER-phagy receptors, which connect the ER to the autophagy machinery. Previous studies have identified different ER-phagy receptors. The involvement of autophagy in the degradation of Golgi however, has not been demonstrated. This thesis present detailed studies of CALCOCO1, and show that it is an autophagy receptor for the degradation of the ER and Golgi apparatus. In the first paper, it is shown that CALCOCO1 is homomeric and that a proportion of the protein localizes in the Golgi apparatus. Functional studies revealed CALCOCO1 is a soluble ER-phagy receptor for the degradation of tubular ER in response to proteotoxic- and starvation-induced stress. On the ER membrane, CALCOCO1 interacts with VAMP-associated proteins VAPA and VAPB via an evolutionary conserved FFAT-like motif and recruits autophagy machinery by binding directly to ATG8 proteins via LIR and UDS interacting region (UIR) motifs acting co-dependently. Depletion of CALCOCO1 caused expansion of the ER and inefficient basal autophagy. In the second paper, involvement of autophagy in the degradation of Golgi apparatus is, for the first time, demonstrated and CALCOCO1 is revealed to be the selective autophagy receptor for the degradation in response to nutrients deprivation. CALCOCO1 interaction with Golgi membrane occurs by binding to the cytoplasmic Ankyrin repeats (AR) domains of membrane-bound Golgi-resident palmitoyltransferases ZDHHC17 and ZDHHC13 via an evolutionary conserved zDHHC-AR-binding motif (zDABM) located at the C-terminal half of the protein. The zDABM motif was also identified and validated in the C-terminal region of the CALCOCO1 paralog, TAX1BP1. Inhibition of autophagy or depletion of CALCOCO1 caused expansion of the Golgi and accumulation of its structural and transmembrane proteins.