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Mol Biol Cell. 2016 Dec 15;27(25):3964-3971. Epub 2016 Oct 26.

Dissecting Torsin/cofactor function at the nuclear envelope: a genetic study.

Author information

1
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520.
2
Department of Cell Biology, Yale School of Medicine, New Haven, CT 06520.
3
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520 christian.schlieker@yale.edu.

Abstract

The human genome encodes four Torsin ATPases, the functions of which are poorly understood. In this study, we use CRISPR/Cas9 engineering to delete all four Torsin ATPases individually and in combination. Using nuclear envelope (NE) blebbing as a phenotypic measure, we establish a direct correlation between the number of inactivated Torsin alleles and the occurrence of omega-shaped herniations within the lumen of the NE. A similar, although not identical, redundancy is observed for LAP1 and LULL1, which serve as regulatory cofactors for a subset of Torsin ATPases. Unexpectedly, deletion of Tor2A in a TorA/B/3A-deficient background results in a stark increase of bleb formation, even though Tor2A does not respond to LAP1/LULL1 stimulation. The robustness of the observed phenotype in Torsin-deficient cells enables a structural analysis via electron microscopy tomography and a compositional analysis via immunogold labeling. Ubiquitin and nucleoporins were identified as distinctively localizing components of the omega-shaped bleb structure. These findings suggest a functional link between the Torsin/cofactor system and NE/nuclear pore complex biogenesis or homeostasis and establish a Torsin-deficient cell line as a valuable experimental platform with which to decipher Torsin function.

PMID:
27798237
PMCID:
PMC5156537
[Available on 2017-03-02]
DOI:
10.1091/mbc.E16-07-0511
[Indexed for MEDLINE]
Free PMC Article

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