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The AAA+ protein torsinA interacts with a conserved domain present in LAP1 and a novel ER protein.

Goodchild RE, Dauer WT - J. Cell Biol. (2005)

Bottom Line: Although the majority of torsinA resides within the endoplasmic reticulum (ER), torsinA binds a substrate in the lumen of the nuclear envelope (NE), and the DeltaE mutation enhances this interaction.Furthermore, we identify a novel transmembrane protein, lumenal domain like LAP1 (LULL1), which also appears to interact with torsinA.Interestingly, LULL1 resides in the main ER.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Columbia University, New York, NY 10032, USA.

ABSTRACT
A glutamic acid deletion (DeltaE) in the AAA+ protein torsinA causes DYT1 dystonia. Although the majority of torsinA resides within the endoplasmic reticulum (ER), torsinA binds a substrate in the lumen of the nuclear envelope (NE), and the DeltaE mutation enhances this interaction. Using a novel cell-based screen, we identify lamina-associated polypeptide 1 (LAP1) as a torsinA-interacting protein. LAP1 may be a torsinA substrate, as expression of the isolated lumenal domain of LAP1 inhibits the NE localization of "substrate trap" EQ-torsinA and EQ-torsinA coimmunoprecipitates with LAP1 to a greater extent than wild-type torsinA. Furthermore, we identify a novel transmembrane protein, lumenal domain like LAP1 (LULL1), which also appears to interact with torsinA. Interestingly, LULL1 resides in the main ER. Consequently, torsinA interacts directly or indirectly with a novel class of transmembrane proteins that are localized in different subdomains of the ER system, either or both of which may play a role in the pathogenesis of DYT1 dystonia.

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TorsinA interacts with the conserved lumenal domain of LAP1. (A) Schematic illustration of LAP1 protein structure and the deletion mutants used in this study. (B) Immunofluorescent labeling of transfected BHKGFPEQ cells with anti-GFP and anti-myc antibodies. GFPEQ-torsinA is displaced by either LAP1 lumenal fragment but not by the lumenal fragment of gp210. (C) Immunofluorescent labeling of BHKGFPΔE cells transfected with myc-tagged 210LAP1 with anti-GFP and anti-myc antibodies.
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fig3: TorsinA interacts with the conserved lumenal domain of LAP1. (A) Schematic illustration of LAP1 protein structure and the deletion mutants used in this study. (B) Immunofluorescent labeling of transfected BHKGFPEQ cells with anti-GFP and anti-myc antibodies. GFPEQ-torsinA is displaced by either LAP1 lumenal fragment but not by the lumenal fragment of gp210. (C) Immunofluorescent labeling of BHKGFPΔE cells transfected with myc-tagged 210LAP1 with anti-GFP and anti-myc antibodies.

Mentions: Next, we examined whether or not the lumenal domain of LAP1 is responsible for its interaction with torsinA, as predicted by our model. To explore this question, we tested if the isolated lumenal domain of LAP1 is capable of altering the perinuclear subcellular distribution of EQ-torsinA. We generated myc-tagged constructs containing the LAP1 lumenal domain with (myc-210LAP1) or without (myc-240LAP1) the transmembrane domain (Fig. 3 A; Kondo et al., 2002). As expected, these fragments fail to concentrate in the NE and instead localize in the main ER (Fig. 3 B, left). Expression of either LAP1 lumenal fragment produced a clear redistribution of GFPEQ-torsinA from the NE to the ER (Fig. 3 B). Myc-210LAP1 causes a similar redistribution of disease-associated GFPΔE-torsinA (Fig. 3 C), and in all instances we observed strong colocalization between labeling for GFP and myc (Fig. 3, B and C). The effect of the LAP1 lumenal domain was specific, as the lumenal domain of the nucleoporin gp210 (Wozniak and Blobel, 1992) did not alter the subcellular distribution of GFPEQ-torsinA (Fig. 3 B, bottom).


The AAA+ protein torsinA interacts with a conserved domain present in LAP1 and a novel ER protein.

Goodchild RE, Dauer WT - J. Cell Biol. (2005)

TorsinA interacts with the conserved lumenal domain of LAP1. (A) Schematic illustration of LAP1 protein structure and the deletion mutants used in this study. (B) Immunofluorescent labeling of transfected BHKGFPEQ cells with anti-GFP and anti-myc antibodies. GFPEQ-torsinA is displaced by either LAP1 lumenal fragment but not by the lumenal fragment of gp210. (C) Immunofluorescent labeling of BHKGFPΔE cells transfected with myc-tagged 210LAP1 with anti-GFP and anti-myc antibodies.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171781&req=5

fig3: TorsinA interacts with the conserved lumenal domain of LAP1. (A) Schematic illustration of LAP1 protein structure and the deletion mutants used in this study. (B) Immunofluorescent labeling of transfected BHKGFPEQ cells with anti-GFP and anti-myc antibodies. GFPEQ-torsinA is displaced by either LAP1 lumenal fragment but not by the lumenal fragment of gp210. (C) Immunofluorescent labeling of BHKGFPΔE cells transfected with myc-tagged 210LAP1 with anti-GFP and anti-myc antibodies.
Mentions: Next, we examined whether or not the lumenal domain of LAP1 is responsible for its interaction with torsinA, as predicted by our model. To explore this question, we tested if the isolated lumenal domain of LAP1 is capable of altering the perinuclear subcellular distribution of EQ-torsinA. We generated myc-tagged constructs containing the LAP1 lumenal domain with (myc-210LAP1) or without (myc-240LAP1) the transmembrane domain (Fig. 3 A; Kondo et al., 2002). As expected, these fragments fail to concentrate in the NE and instead localize in the main ER (Fig. 3 B, left). Expression of either LAP1 lumenal fragment produced a clear redistribution of GFPEQ-torsinA from the NE to the ER (Fig. 3 B). Myc-210LAP1 causes a similar redistribution of disease-associated GFPΔE-torsinA (Fig. 3 C), and in all instances we observed strong colocalization between labeling for GFP and myc (Fig. 3, B and C). The effect of the LAP1 lumenal domain was specific, as the lumenal domain of the nucleoporin gp210 (Wozniak and Blobel, 1992) did not alter the subcellular distribution of GFPEQ-torsinA (Fig. 3 B, bottom).

Bottom Line: Although the majority of torsinA resides within the endoplasmic reticulum (ER), torsinA binds a substrate in the lumen of the nuclear envelope (NE), and the DeltaE mutation enhances this interaction.Furthermore, we identify a novel transmembrane protein, lumenal domain like LAP1 (LULL1), which also appears to interact with torsinA.Interestingly, LULL1 resides in the main ER.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Columbia University, New York, NY 10032, USA.

ABSTRACT
A glutamic acid deletion (DeltaE) in the AAA+ protein torsinA causes DYT1 dystonia. Although the majority of torsinA resides within the endoplasmic reticulum (ER), torsinA binds a substrate in the lumen of the nuclear envelope (NE), and the DeltaE mutation enhances this interaction. Using a novel cell-based screen, we identify lamina-associated polypeptide 1 (LAP1) as a torsinA-interacting protein. LAP1 may be a torsinA substrate, as expression of the isolated lumenal domain of LAP1 inhibits the NE localization of "substrate trap" EQ-torsinA and EQ-torsinA coimmunoprecipitates with LAP1 to a greater extent than wild-type torsinA. Furthermore, we identify a novel transmembrane protein, lumenal domain like LAP1 (LULL1), which also appears to interact with torsinA. Interestingly, LULL1 resides in the main ER. Consequently, torsinA interacts directly or indirectly with a novel class of transmembrane proteins that are localized in different subdomains of the ER system, either or both of which may play a role in the pathogenesis of DYT1 dystonia.

Show MeSH
Related in: MedlinePlus