The effects of different neurodegenerative diseases on neurons substantially differ, but the accumulation of intracellular toxic aggregates of misfolded proteins and the consequent loss of cell function are common features of all neurodegenerative diseases. The underlying reason for the presence of aggregates is not known in the vast majority of cases, but some genes mutated in neurodegenerative diseases are linked to autophagy. Autophagy is an evolutionarily conserved process by which noxious cytoplasmic contents such as protein aggregates, pathogens, and damaged organelles are broken down in lysosomes. Autophagy adaptors recognize the cargo labeled for degradation and deliver it to double-membrane vesicles called autophagosomes. Autophagosomes fuse with lysosomes leading to the formation of autolysosomes where the cargo is degraded. Autophagy adapters also include two proteins whose mutations have been found in patients with the neurodegenerative disease amyotrophic lateral sclerosis (ALS), optineurin (OPTN) and sequestosome 1 (SQSTM1 or p62). In addition to its adaptor role, OPTN has also been described to participate in two other autophagy steps: autophagy initiation and autophagosome maturation. Therefore, in this paper, we examined the effect of OPTN on autophagy in the N2a neuroblastoma cell line, using N2a cells in which OPTN was removed by the CRISPR-Cas9 approach. We analyzed basal autophagy and autophagy induced by interferon (IFN)-, a cytokine recently shown to regulate autophagy in neurons. We used standard tests to monitor autophagy such as measuring the amount of LC3-II (microtubule associated protein light chain 3), p62 adaptor, and OPTN by Western blotting, and counting LC3 positive vesicles by immunofluorescence or via a fluorescently labeled LC3 construct. We observed a partial block of basal autophagy in the OPTN-deficient cells manifested as an increased number and size of autophagosomes and increased accumulation of LC3-II. IFN-β induced autophagy in N2a cells, but we did not observe the accumulation of OPTN and p62 which may mean that they do not serve as autophagy adaptors upon IFN-β-treatment or that they act on autophagy in some other step(s). Furthermore, it is possible that they were degraded before our analysis and that our experimental approach needs to be modified. Due to the partial block in autophagy in the absence of OPTN, it is possible that this leads to an accumulation of protein aggregates in ALS. In future experiments, it is therefore important to test if this observed autophagy dysregulation in OPTN-deficient cells leads to protein aggregation.