ROCK-phosphorylated vimentin modifies mutant huntingtin aggregation via sequestration of IRBIT

Peter O Bauer 1 Roman Hudec 0 Anand Goswami 1 Masaru Kurosawa 1 Gen Matsumoto 1 Katsuhiko Mikoshiba 0 Nobuyuki Nukina 1 0 Laboratory for Developmental Neurobiology, Brain Science Institute, RIKEN , 2-1 Hirosawa, Wako-shi, Saitama 351-0198 , Japan 1 Laboratory for Structural Neuropathology, Brain Science Institute, RIKEN , 2-1 Hirosawa, Wako-shi, Saitama 351-0198 , Japan Background: Huntington's Disease (HD) is a fatal hereditary neurodegenerative disease caused by the accumulation of mutant huntingtin protein (Htt) containing an expanded polyglutamine (polyQ) tract. Activation of the channel responsible for the inositol-induced Ca2+ release from ensoplasmic reticulum (ER), was found to contribute substantially to neurodegeneration in HD. Importantly, chemical and genetic inhibition of inositol 1,4,5-trisphosphate (IP3) receptor type 1 (IP3R1) has been shown to reduce mutant Htt aggregation. Results: In this study, we propose a novel regulatory mechanism of IP3R1 activity by type III intermediate filament vimentin which sequesters the negative regulator of IP3R1, IRBIT, into perinuclear inclusions, and reduces its interaction with IP3R1 resulting in promotion of mutant Htt aggregation. Proteasome inhibitor MG132, which causes polyQ proteins accumulation and aggregation, enhanced the sequestration of IRBIT. Furthermore we found that IRBIT sequestration can be prevented by a rho kinase inhibitor, Y-27632. Conclusions: Our results suggest that vimentin represents a novel and additional target for the therapy of polyQ diseases. - Background Huntington's disease (HD) is an autosomal-dominant neurodegenerative disorder caused by CAG repeat expansion coding for a polyglutamine (polyQ) sequence in the N-terminal region of the huntingtin protein (Htt). The expansion of more than 36 repeats causes misfolding of the gene product huntingtin resulting in a toxic gain-of-function [1]. Clinically, HD is characterized by chronic and progressive involuntary choreiform movements, mood disorders, cognitive impairment, and behavioral changes [2,3]. A prominent feature of this disease is progressive neurodegeneration, with neuronal intranuclear and cytoplasmic accumulation of aggregated polyQ protein [4,5]. HD pathomechanism involves a broad scale of events including dysregulation of transcription and gene expression, impairment of axonal transport and synaptic transmission and impairment of the ubiquitin proteasome system (UPS) [6,7]. Mitochondrial dysfunction leading to induction of mitochondrial apoptotic pathway has also been described in HD with Ca2+ mishandling and suppression of energy metabolism [8,9]. Despite an enormous effort in elucidating the pathogenesis of this disorder, effective therapies for HD have not yet been found. Vimentin is a 57 kDa type III intermediate filament (IF) found in cells of mesenchymal origin [10,11]. While widely expressed in embryos, vimentin is replaced by other major classes of IFs in cells during terminal differentiation [12,13]. In the adult brain, vimentin expression is mostly restricted to some subpopulations of glial and vascular endothelial cells under physiological conditions [12-14]. Importantly, it has been found that vimentin expression is re-activated in mature neurons affected by Alzheimers disease or traumatic injury [15,16]. Degradation of misfolded proteins has been shown partly mediated by UPS. The components of UPS including the 26S proteasome and ubiquitin as well as heat shock proteins are concentrated at the centrosome [17]. When UPS is overloaded by misfolded proteins and/or it is chemically inhibited, the centromeric accumulation of these proteins increases forming aggresomes which may represent a general cellular response to dysfunctional or damaged polyubiquitinated proteins accumulation [18,19]. Another evidence of the association of aggresome formation with the accumulation and degradation of misfolded proteins has come from studies, where pathogenic polyQ proteins Htt and atrophin-1 formed inclusions at centrosomes which were surrounded by vimentin [20,21]. Vimentin is recruited to the aggresomes during UPS dysfunction and forms a cage-like structure surrounding the pericentriolar focus of aggregated protein [19]. The role of aggresomes and especially the vimentin cage in polyQ diseases progression is not clear. We hypothesized that vimentin may play a major role in polyQ proteins accumulation and aggregation and that vimentin cage may immobilize or trap not only the UPS components and chaperones, but also other important proteins at the centrosomic inclusions and thus preventing their functions elsewhere in the cell. We found that one of such proteins, IP3R1-interacting protein released with IP3 (IRBIT) is also sequestered by vimentin. IRBIT has numerous regulatory functions among which the IP3R1 activity regulation is most intriguing [22,23]. IRBIT binds to IP3-binding core domain of IP3R1 acting as a competitor to IP3 [23,24]. Absence of IRBIT sen (...truncated)