Pathological consequences of VCP-mutations on human striated muscle: Nuclear and cytoplasmic inclusions, a distinct form of dilatative cardiomyopathy, and a novel link to VCP-glycoprotein interactions

Abstract

Mutations in the valosin containing protein (VCP, p97) gene cause a late-onset form of autosomal dominant Inclusion Body Myopathy associated with Paget disease of the bone and Frontotemporal Dementia (IBMPFD). VCP is an ubiquitously expressed member of the AAA-ATPase family with a tripartite structure comprising an N-terminal domain (Cdc48) involved in ubiquitin-binding, and two central D1- and D2-domains which hydrolyse ATP. VCP assembles into functional hexamers with a central cylinder formed by the D-domains surrounded by the N-domains. VCP has been associated with a wide variety of essential cellular processes including the ubiquitin proteasome protein degradation system and along with its co-factors Udf1, Npl4, and Derlin-1 with the endoplasmic reticulum associated protein degradation (ERAD). We report on the pathological consequences of three heterozygous VCP-mutations (R93C, R155H, R155C; located in the N-terminal Cdc48 domain of the VCP protein) on human striated muscle in vivo and in vitro. IBMPFD skeletal muscle pathology is characterized by degenerative changes and filamentous VCP- and ubiquitin-positive cytoplasmic and nuclear protein aggregates. Furthermore, mutant VCP leads to a novel form of dilatative cardiomyopathy with inclusion bodies. In contrast to post-mitotic striated muscle cells protein aggregate pathology was neither detected in primary IBMPFD myoblasts nor in transiently and stably transfected cells using wildtype-VCP and the VCP mutants. Pull-down experiments showed that all three VCP mutations do not affect the binding to Ufd1, Npl4, and ataxin-3. Structural analysis demonstrated that R93 and R155 are both surface-accessible residues located in the center of cavities that may enable ligand binding. Mutations at R93 and R155 are predicted to induce changes in the tertiary structure of the VCP protein. The search for putative ligands to the R93 and R155 cavities resulted in the identification of cyclic sugar compounds providing a novel link to VCP carbohydrate interactions in the complex pathology of IBMPFD.