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

Abstract

TorsinA (also known as torsin-1A) is a membrane-embedded AAA+ ATPase that has an important role in the nuclear envelope lumen. However, most torsinA is localized in the peripheral endoplasmic reticulum (ER) lumen where it has a slow mobility that is incompatible with free equilibration between ER subdomains. We now find that nuclear-envelope-localized torsinA is present on the inner nuclear membrane (INM) and ask how torsinA reaches this subdomain. The ER system contains two transmembrane proteins, LAP1 and LULL1 (also known as TOR1AIP1 and TOR1AIP2, respectively), that reversibly co-assemble with and activate torsinA. Whereas LAP1 localizes on the INM, we show that LULL1 is in the peripheral ER and does not enter the INM. Paradoxically, interaction between torsinA and LULL1 in the ER targets torsinA to the INM. Native gel electrophoresis reveals torsinA oligomeric complexes that are destabilized by LULL1. Mutations in torsinA or LULL1 that inhibit ATPase activity reduce the access of torsinA to the INM. Furthermore, although LULL1 binds torsinA in the ER lumen, its effect on torsinA localization requires cytosolic-domain-mediated oligomerization. These data suggest that LULL1 oligomerizes to engage and transiently disassemble torsinA oligomers, and is thereby positioned to transduce cytoplasmic signals to the INM through torsinA.

Sequences within the C Terminus of the Metabotropic Glutamate Receptor 5 (mGluR5) Are Responsible for Inner Nuclear Membrane Localization

“…Immunostained samples were quantitated in a blind fashion such that regions from the plasma membrane and nuclear membrane were selected by using transmitted light images and lamin B 2 images, respectively, and the ratio of nuclear membrane to the plasma membrane was compared between different constructs. Line scan analysis was performed by randomly drawing a horizontal line near the center of the nucleus followed by a vertical line perpendicular to the first (73). Fluorescence intensity was quantified along both lines using MetaMorph software.…”

Torsins: not your typical AAA+ ATPases

“…Overexpression of LULL1 leads to a redistribution of TorsinA WT from the ER to the NE, but Walker A (K108A) and Walker B (E171Q) mutants as well as TorsinA lacking its hydrophobic domain are not affected (Vander Heyden et al, 2009, Goodchild et al, 2015). This suggests that LULL1 may play a role in regulating the subcellular distribution of TorsinA.…”

“…In fact, it has been previously shown that mutation of cysteines in TorsinA affects both nucleotide and cofactor binding (Zhu et al, 2008, Zhu et al, 2010). LAP1 forms a complex with TorsinA in vitro (Brown et al, 2014, Zhao et al, 2013) (Figure 5A), but in vivo conditions (or crosslinking in vitro ) are required to observe higher-order oligomers (Jungwirth et al, 2010, Vander Heyden et al, 2009, Li et al, 2014, Brown et al, 2014, Goodchild et al, 2015). The precise composition of these oligomers remains poorly understood.…”

“…Binding of interaction partners to LAP1 and LULL1 or their modification could induce conformational changes that propagate across the membrane to regulate Torsin activity on the luminal side (Brown et al, 2014). Indeed, recent evidence suggests that the cytosolic domain of LULL1 oligomerizes and that its oligomerization state affects the stoichiometry of the overall assembly as well as its subcellular localization (Goodchild et al, 2015). Therefore, only a thorough characterization of full-length components in the natural context of the membrane (e.g.…”

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

“…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).…”