ATAD3 proteins in autophagy. Do ATAD3 proteins have a direct role in regulating autophagy

Abstract

ATPase Family AAA Domain -containing protein 3 (ATAD3) is a nuclear-encoded mitochondrial transmembrane protein encompassing three isoforms; ATAD3A, ATAD3B and ATAD3C. ATAD3 is involved in several cellular process and available data predict that ATAD3A and ATAD3B proteins may have potential roles in regulating autophagy and mitophagy. Preliminary data from our research group identified interactions between ATAD3A and ATAD3B with ATG8 proteins, p62, NDP52 and CALCOCO1. However, whether these interactions play any role in autophagy is not known. This study sought to characterise these interactions and evaluate their roles in autophagy or mitophagy. In vitro GST pulldown analysis together with autoradiography were used to probe the interaction between ATAD3 proteins and some autophagy-related proteins including LC3B, GABARAP, p62, NDP52 and CALCOCO1. Western blot analysis was employed to evaluate ATAD3A levels following starvation-induced autophagy, while immunostaining and confocal microscopy were used to assess ATAD3 colocalization with p62 and LC3B. Results showed that interaction between ATAD3 and LC3B and GABARAP is not mediated by a LIR motif. Instead, the interaction is dependent on the C-terminal ATPase domain. Likewise, the interaction with the autophagic receptors: p62, NDP52 and CALCOCO1 is mediated by the C-terminal region which has been reported to be embedded in the mitochondrial matrix. Moreover, ATAD3A is not degraded by starvation-induced autophagy; rather, its translation is upregulated.