The mechanism of Torsin ATPase activation

Abstract

Torsins are membrane-associated ATPases whose activity is dependent on two activating cofactors, lamina-associated polypeptide 1 (LAP1) and luminal domain-like LAP1 (LULL1). The mechanism by which these cofactors regulate Torsin activity has so far remained elusive. In this study, we identify a conserved domain in these activators that is predicted to adopt a fold resembling an AAA+ (ATPase associated with a variety of cellular activities) domain. Within these domains, a strictly conserved Arg residue present in both activating cofactors, but notably missing in Torsins, aligns with a key catalytic Arg found in AAA+ proteins. We demonstrate that cofactors and Torsins associate to form heterooligomeric assemblies with a defined Torsin-activator interface. In this arrangement, the highly conserved Arg residue present in either cofactor comes into close proximity with the nucleotide bound in the neighboring Torsin subunit. Because this invariant Arg is strictly required to stimulate Torsin ATPase activity but is dispensable for Torsin binding, we propose that LAP1 and LULL1 regulate Torsin ATPase activity through an active site complementation mechanism.DYT1 dystonia | nuclear envelope | ATPase | Torsin T orsin ATPases belong to the AAA+ (ATPase associated with a variety of cellular activities) superfamily of ATPases (1) but are unusual in that they lack conserved catalytic residues that are typically found in related ATPases (2, 3). Accordingly, Torsins do not display ATPase activity unless they are engaged by their regulatory cofactors lamina-associated polypeptide 1 (LAP1) or luminal domain-like LAP1 (LULL1) (4), which are type II transmembrane proteins residing in the nuclear envelope and endoplasmic reticulum (ER) (5, 6). This property stands in sharp contrast to the behavior of closely related Clp/Hsp100 ATPases, which display considerable basal ATPase activities that are moderately stimulated by their substrates (7,8). Although a number of Clp/Hsp100 proteins use distinct cofactors that confer substrate specificity to the typically hexameric ATPase ring, they operate via similar principles (9). Substrates ultimately engage the pore at the center of the oligomeric assembly. This narrow annulus is defined by pore loops that emanate from each subunit and harbor a conserved aromatic residue that defines the center of the pore (10). The energy of ATP hydrolysis is invested in threading the substrate through this axial channel, and the substrate is unfolded in the process (11).Much less is known about the mode of action of Torsin ATPases, although it is clear that regulation of Torsin activity is of vital importance in an organismal context. A mutation in TorsinA (TorA) causing the movement disorder primary dystonia (12) renders TorA unresponsive to its binding partners LAP1 and LULL1 (4), and a homozygous "knock-in" of this disease allele in mice causes a lethal phenotype, as does a TorA KO (13). A conditional deletion of TorA from the CNS in mice accurately replicates the symptoms of primary dystonia (14). In add...

Access of torsinA to the inner nuclear membrane is activity dependent and regulated in the endoplasmic reticulum

“…Most, including torsinA, are embedded in the luminal leaflet of the endoplasmic reticulum (ER) membrane (Jungwirth et al, 2010;Vander Heyden et al, 2011). Importantly, torsinA binds two transmembrane proteins of the ER system, LAP1 (also known as TOR1AIP1) and LULL1 (also known as TOR1AIP2 and NET9) Kim et al, 2010;Naismith et al, 2009;Zhao et al, 2013;Zhu et al, 2010), that have recently been shown to adopt a partial AAA+ domain fold and co-assemble with torsinA as subunits of an active enzyme (Brown et al, 2014;Sosa et al, 2014). The luminal domains of LAP1 and LULL1 are homologous to each other, whereas the extraluminal domains diverge .…”

“…The luminal domains of LAP1 and LULL1 are homologous to each other, whereas the extraluminal domains diverge . Like torsins, proteins similar to LAP1 and LULL1 have been identified across animal phyla and might have co-evolved to function together with torsins (Brown et al, 2014;Sosa et al, 2014).…”

Site-specific Proteolysis Mobilizes TorsinA from the Membrane of the Endoplasmic Reticulum (ER) in Response to ER Stress and B Cell Stimulation

“…1A and 6A), we consider a major role in counteracting protein misfolding unlikely. Interestingly, several of the Torsins (Tor2A and Tor3A in humans and mice) do not harbor a hydrophobic domain, and TorA variants deprived of their hydrophobic domains retain ATPase activity (16,17), suggesting that mobilized TorAp could have a function that is distinct from its precursor.…”

“…TorsinA is partitioned between the ER and the contiguous perinuclear space of the nuclear envelope, and its localization is controlled in part by its association with LAP1 and LULL1, type II transmembrane proteins residing in the nuclear envelope and ER, respectively (12)(13)(14). Another unusual property of Torsins is that they lack significant basal ATPase activity in isolation and are tightly regulated by LAP1 and LULL1, which integrate into the Torsin ring via AAA-like domains to trigger ATP hydrolysis through an active site complementation mechanism (15)(16)(17). Thus, Torsins and their ATPase activating cofactors form a composite, membrane-spanning assembly.…”

“…Thus, Torsins and their ATPase activating cofactors form a composite, membrane-spanning assembly. This assembly is sensitive to the redox environment of the ER (18) and is perturbed by the DYT1 dystonia-causing mutation, presumably due to a defect in the Torsin-activator interface (16), underscoring the importance of the regulation of this assembly. Although a key role for a nucleotide-proximal cysteine has been established in the context of redox regulation (18,19), the functional significance of the cysteine cluster positioned near the luminal membrane interface is presently unclear.…”