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Autophagy is a mechanism responsible for intracellular degradation and recycling of macromolecules and organelles, essential for cell survival in adverse conditions. More than 40 autophagy-related ( ATG ) genes have been identified and characterized in fungi, among them  ATG4  and  ATG8 .  ATG4  encodes a cysteine protease (Atg4) that plays an important role in autophagy by initially processing Atg8 at its C-terminus region. Atg8 is a ubiquitin-like protein essential for the synthesis of the double-layer membrane that constitutes the autophagosome vesicle, responsible for delivering the cargo from the cytoplasm to the vacuole lumen. The contributions of Atg-related proteins in the pathogenic yeast in the genus  Cryptococcus  remain to be explored, to elucidate the molecular basis of the autophagy pathway. In this context, we aimed to investigate the role of autophagy-related proteins 4 and 8 (Atg4 and Atg8) during autophagy induction and their contribution with non-autophagic events in  C .  neoformans . We found that Atg4 and Atg8 are conserved proteins and that they interact physically with each other.  ATG  gene deletions resulted in cells sensitive to nitrogen starvation.  ATG4  gene disruption affects Atg8 degradation and its translocation to the vacuole lumen, after autophagy induction. Both  atg4  and  atg8  mutants are more resistant to oxidative stress, have an impaired growth in the presence of the cell wall-perturbing agent Congo Red, and are sensitive to the proteasome inhibitor bortezomib (BTZ). By that, we conclude that in  C .  neoformans  the autophagy-related proteins Atg4 and Atg8 play an important role in the autophagy pathway; which are required for autophagy regulation, maintenance of amino acid levels and cell adaptation to stressful conditions.

Biological functions of the autophagy-related proteins Atg4 and Atg8 in Cryptococcus neoformans