Inflammation in Parkinson's disease (PD) is tightly associated with disease pathogenesis. The mutations in the Omi (PARK13) gene, which encodes the protease Omi/HtrA2, are linked to neurodegeneration and PD. Loss of Omi protease activity by the point mutation S276C leads to severe neurodegeneration and neuroinflammation in the striatum and spinal cord in mnd2 (motor neuron degeneration 2) mice. We show that Omi plays roles in the activation of microglia. We identified that MEK1 (mitogen-activated protein kinase kinase 1) is a substrate of Omi. Omi cleaves MEK1, to suppress its downstream kinase ERK1/2 (Extracellular Signal-Regulated Kinase 1/2) activation, thereby regulating the expression of microglial inflammatory molecules. Knockdown of Omi in microglial cells activates ERK1/2 and results in IκBα (inhibitor of NF-κB) degradation, which induces the expression of inflammatory molecules, such as tumor necrosis factor (TNF-α) and inducible nitric oxide synthase (iNOS). The expression of inflammatory molecules induced by the knockdown of Omi can be blocked by U0126, a specific inhibitor of MEK1, suggesting an involvement of MAPK signaling in Omi-deficiency-induced microglial activation. Furthermore, the S276C Omi mutant, which is a protease-deficient mutant, has no effect on MEK1 cleavage or ERK1/2 activation. In the brains of mnd2 mice that carry the S276C Omi mutation, increased transcription of several inflammatory molecules and the activation of astrocytes and microglia are observed. Therefore, our study demonstrates that Omi is an intrinsic factor in the regulation of neuroinflammation.