Role of ALS-associated OPTN-K489E mutation in neuronal cell-death regulation.

Optineurin (OPTN) gene is a marker of amyotrophic lateral sclerosis (ALS). However, the role of optineurin protein (OPTN) in ALS pathology is unclear, even though it is known to regulate autophagy, apoptosis, and other survival-death cellular processes. Genetic analysis of Indian ALS patients by our group ascertained a novel mutation K489E in the OPTN gene. To identify the molecular mechanism associated with OPTN and its mutation, we developed an in-vitro cell model using SH-SY5Y cells harbouring OPTN and OPTN-K489E mutation along with its control vector. Since we observed a significant decrease in cell viability in the mutant, we measured the expressions of genes and proteins mediating apoptosis, necroptosis, and autophagy, to establish the role of OPTN in cell death regulation. Our results show that OPTN-K489E mutation changes the relative gene expressions of miRNA-9, REST, CoREST and BDNF, and causes apoptosis. We also observed an up-regulation in the expressions of necroptosis mediated genes RIPK1, RIPK3, and MLKL and autophagy mediated genes TBK1, P62, and LC3II. The results of FACS analyses revealed that this mutation promotes apoptotic and necroptotic processes confirming the pathogenicity of OPTN-K489E.