Knockdown of TMEM160 leads to an increase in reactive oxygen species generation and the induction of the mitochondrial unfolded protein response

Transmembrane protein 160 (TMEM160) was recently reported to be localized to the mitochondrial inner membrane, but mitochondrial function was noted to be unaffected by loss of TMEM160. In contrast to these previously published findings, we report here that the absence of TMEM160 influences intracellular responses. After confirming that TMEM160 is localized in the inner mitochondrial membrane, we knocked down TMEM160 in human cultured cells and analyzed the changes in cellular responses. TMEM160 depletion led to an upregulation of the mitochondrial chaperone HSPD1, suggesting that depletion induced the mitochondrial unfolded protein response (UPR(mt)). Indeed, the expression of key transcription factors that induce the UPR(mt) (ATF4, ATF5, and DDIT3) was increased following TMEM160 depletion. Expression of the mitochondrial protein import‐receptors TOMM22 and TOMM20 was also enhanced. In addition, we observed a significant increase in reactive oxygen species (ROS) generation following TMEM160 depletion. Glutathione S‐transferases, which detoxify the products of oxidative stress, were also upregulated in TMEM160‐depleted cells. Immunoblot analysis was performed to detect proteins modified by 4‐hydroxynonenal (which is released after the peroxidation of lipids by ROS): the expression patterns of 4‐hydroxynonenal‐modified proteins were altered after TMEM160 depletion, suggesting that depletion enhanced degradation of these proteins. HSPD1, TOMM22, ATF4, ATF5, and DDIT3 remained upregulated after ROS was scavenged by N‐acetylcysteine, suggesting that once the UPR(mt) is induced by TMEM160 depletion, it is not suppressed by the subsequent detoxification of ROS. These findings suggest that TMEM160 may suppress ROS generation and stabilize mitochondrial protein(s).