Abstract
BackgroundDYT1 dystonia is an autosomal dominant neurological condition caused by a mutation that removes a single glutamic acid residue (ΔE) from the torsinA (torA) AAA+ protein. TorA appears to possess a nuclear envelope (NE) localized activity that requires Lamina-Associated-Polypeptide 1 (LAP1), which is an inner nuclear membrane localized torA-binding partner. Although hypoactive, the DYT1 dystonia torA-ΔE isoform often concentrates in the NE, suggesting that torA-ΔE also interacts with an NE-localized binding partner.ResultsWe confirm that NE-localized torA-ΔE does not co-immunoprecipitate with LAP1, and find that torA-ΔE continues to concentrate in the NE of cells that lack LAP1. Instead, we find that variability in torA-ΔE localization correlates with the presence of the SUN-domain and Nesprin proteins that assemble into the LINC complex. We also find that siRNA depletion of SUN1, but not other LINC complex components, removes torA-ΔE from the NE. In contrast, the LAP1-dependent NE-accumulation of an ATP-locked torA mutant is unaffected by loss of LINC complex proteins. This SUN1 dependent torA-ΔE localization requires the torA membrane association domain, as well as a putative substrate-interaction residue, Y147, neither of which are required for torA interaction with LAP1. We also find that mutation of these motifs, or depletion of SUN1, decreases the amount of torA-WT that colocalizes with NE markers, indicating that each also underlies a normal NE-localized torA binding interaction.ConclusionsThese data suggest that the disease causing ΔE mutation promotes an association between torA and SUN1 that is distinct to the interaction between LAP1 and ATP-bound torA. This evidence for two NE-localized binding partners suggests that torA may act on multiple substrates and/or possesses regulatory co-factor partners. In addition, finding that the DYT1 mutation causes abnormal association with SUN1 implicates LINC complex dysfunction in DYT1 dystonia pathogenesis, and suggests a gain-of-function activity contributes to this dominantly inherited disease.
Highlights
DYT1 dystonia is an autosomal dominant neurological condition caused by a mutation that removes a single glutamic acid residue (ΔE) from the torsinA AAA+ protein
TorA is a luminal protein, torA-E171Q and torA-ΔE concentrate in the nuclear envelope (NE) of several cell lines, and each diffuses more slowly than endoplasmic reticulum (ER)-localized torA [12], which suggests that both isoforms interact with a lamina-associated NE membrane protein, such as Lamina-Associated-Polypeptide 1 (LAP1)
We examined the LAP1-binding of human torA-ΔE and torA-E171Q using mouse NIH-3T3 cells where both isoforms localize in the NE (Figure 1A), whereas this is previously assessed using cell lines where torA-ΔE is predominantly ER-localized [15,20]
Summary
DYT1 dystonia is an autosomal dominant neurological condition caused by a mutation that removes a single glutamic acid residue (ΔE) from the torsinA (torA) AAA+ protein. The disease is caused by an inframe, loss-of-function mutation that removes a glutamic acid residue (ΔE) from torA [3,4]. In most cases, an oligomeric ring arrangement of AAA+ enzyme subunits pulls the binding-partner substrate into the central pore and, by doing so, ‘stretches’ or removes secondary structure from the substrate. This action often destabilizes an otherwise energetically favorable binding interaction, such as the presence of substrate in a protein complex, aggregate, or association with a lipid bilayer. There are many hundreds of AAA+ enzymes and substrates, and AAA+ enzymes are used in processes as diverse as DNA replication, membrane fusion, protein degradation and cytoskeletal movement [5,6,7]
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