Characterization of torsin -related protein 2A: A novel AAA+ chaperone protein

Abstract

Dystonia is a neurologic movement disorder characterized by involuntary twisting movements and abnormal postures. A GAG deletion in TOR1A correlates with the disease early onset dystonia (EOD). Inheritance of EOD is autosomal-dominant, with reduced penetrance (30--40%).;The TOR1A gene family consists of four homologous human genes. Though the protein encoded by TOR1A, torsinA, is part of the AAA+ superfamily of chaperone proteins, which are frequently involved in protein folding and re-folding, the function of the torsin protein family remains elusive. We cloned and characterized TOR2A, another member of the gene family that encodes the protein, torp2a. Torp2a shares 43% amino acid identity with torsinA, and both contain ATPase domains that are homologous to those of the ATP dependent protease family of chaperone proteins.;RNA expression analysis demonstrated that TOR2A mRNA is ubiquitously expressed in human adult and fetal tissues. Furthermore, in situ hybridization of TOR2A mRNA revealed that several regions of the mouse brain displayed strong TOR2A expression. However, none of these areas control movement or have been implicated in movement disorders.;In order to elucidate torp2a's function, its subcellular localization was examined. Immunofluorescence studies demonstrated that torp2a localizes to the endoplasmic reticulum, and co-localizes with the ER resident protein, protein disulfide isomerase. Subcellular fractionation and detergent phase partitioning of torp2a revealed that torp2a predominantly localizes to the ER, but a small fraction of the protein remains in the cytosol. Thus, we conclude that like torsinA, torp2a, is a dually localized protein.;Mass spectrometry was used to identify components of torp2a immunocomplexes. Two proteins, heat shock cognate protein 70 and heat shock protein 90a directly interact with torp2a, which suggested that torp2a functions as a chaperone protein. To determine whether torp2a can re-fold protein aggregates, TOR2A was used to transduce C. elegans lines expressing polyglutamine aggregates. Transduced worms showed significant reduction in the numbers and sizes of protein aggregates, compared to control worms. These results show that torp2a functions as part of a large chaperone complex that mediates protein re-folding in vivo. Understanding the function of torp2a is important to elucidating the physiological function of the torsin protein family, and broadens our understanding of the pathophysiology of EOD.