Actin was used as a loading control. protein 1 (UBXD1), a known co-factor of VCP assisting in the recognition of substrates for protein degradation, selectively binds to MCL1 Rabbit Polyclonal to SFRS4 and interacts with VCP to mediate MCL1 extraction from the mitochondria. These results indicate that the OMM protein MCL1 is degraded by the VCP-UBXD1 complex and that the process is promoted by the presence of mtHtt. Therefore, our finding provides a new insight into the mechanism of mitochondrial dysfunction in HD. Introduction Huntingtons disease (HD) is a fatal, inherited neurodegenerative disorder that progresses for 15C20 years after diagnosis. The mutation that causes HD is a variable expansion of CAG repeats encoding polyglutamine (polyQ) in the huntingtin (Htt) protein (mutant Huntingtin, Finasteride acetate mtHtt) [1]. Many studies, including ours, Finasteride acetate show that mitochondrial dysfunction is one of the major events in the pathogenesis of HD [2C6]. Mitochondria isolated from HD patients or HD-transgenic mouse brains show an increase in mitochondrial permeability, perturbation in mitochondrial membrane potential, changes in mitochondrial ultrastructure and DNA integrity, as well as failure in bio-energetics [7C10]. Moreover, incubation of isolated mitochondria with recombinant mtHtt protein leads to mitochondrial swelling and release of cytochrome c [10, 11], suggesting direct mitochondrial injury by mtHtt. Further, we and others recently demonstrated the impairment of mitochondrial fission and mitophagy in various experimental models of HD [3, 5, 6, Finasteride acetate 12, 13] and that pharmacological inhibition of either mitochondrial fragmentation or aberrant mitophagy reduces neuropathological and behavioral phenotypes in HD transgenic mice [5, 6]. Thus, proper control of mitochondrial quality is likely to be a useful strategy for halting or slowing the pathology of HD. To ensure proper mitochondrial function, diverse mitochondrial quality control mechanisms are evolutionarily conserved. In addition to mitochondrial dynamics (fusion and fission) that are critical for maintaining mitochondrial morphology and transport [12, 14], mitochondria-associated protein degradation (MAD) that occurs at the compartment of the outer mitochondrial membrane (OMM) has been proposed to be one of the mechanisms for controlling normal mitochondrial function [15, 16]. The MAD mechanism involves retro-translocation of ubiquitinated proteins from the OMM to the cytosol where the proteins can be degraded by the ubiquitin-proteasome pathway (UPS) [15, 17, 18]. Valosin-containing-protein (VCP), also known as p97, is an AAA-ATPase central Finasteride acetate to UPS-dependent protein turnover [19]. VCP uses energy derived from ATP hydrolysis to apply mechanical force to substrates, thereby extracting substrates from diverse cellular locations for proteasomal degradation [19, 20]. Several studies reported that VCP translocates to the mitochondria, where it targets ubiquitinated OMM proteins (substrates) with relatively short half-lives before retro-translocating them to the proteasome for degradation [21C24]. We recently found that VCP is recruited to the mitochondria in various experimental models of HD and that blocking VCP mitochondrial translocation by a novel inhibitor reduces neuropathology of HD [5], indicating the importance of mitochondrial VCP in HD. However, whether mitochondria-accumulated VCP is involved in the MAD mechanism in HD is unknown. Myeloid cell leukemia sequence 1 (MCL1) is a nucleus-encoded protein localized both on the OMM and in the mitochondrial matrix [5]. MCL1 is essential for maintaining mitochondrial membrane potential and is required for matrix and inner membrane structure integrity and mitochondrial bioenergetics [25, 26]. MCL1 can also serve as a stress sensor that regulates autophagy [27]. The protein level of MCL1 is controlled by the UPS. Degradation of MCL1 in cells results in extensive mitochondrial damage.