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TorsinA is essential for the timing and localization of neuronal nuclear pore complex biogenesis
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Apr 27, 2023
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1
TorsinA is essential for the timing and localization of neuronal
nuclear pore complex biogenesis
Sumin Kim1,2, Sébastien Phan3, Thomas R. Shaw4, Mark H. Ellisman3, Sarah L. Veatch4, Sami J. Barmada1,2,*,
Samuel S. Pappas5,6,*, and William T. Dauer5,6,7,*
1Cellular and Molecular Biology Graduate Program, University of Michigan, Ann Arbor, MI; 2Department of Neurology, University of Michigan, Ann Arbor,
MI; 3National Center for Microscopy and Imaging Research, Center for Research on Biological Systems, Department of Neurosciences, School of
Medicine, University of California San Diego, La Jolla, CA; 4Department of Biophysics, University of Michigan, Ann Arbor, MI; 5Peter O9Donnell Jr. Brain
Institute, UT Southwestern, Dallas, TX; 6Department of Neurology, UT Southwestern, Dallas, TX; 7Department of Neuroscience, UT Southwestern,
Dallas, TX;
*To whom correspondence should be addressed: sbarmada@med.umich.edu; Samuel.Pappas@utsouthwestern.edu; William.Dauer@utsouthwestern.edu
Nuclear pore complexes (NPCs) regulate information transfer between the nucleus and cytoplasm. NPC
defects are linked to several neurological diseases, but the processes governing NPC biogenesis and
spatial organization are poorly understood. Here, we identify a temporal window of strongly upregulated
NPC biogenesis during neuronal maturation. We demonstrate that the AAA+ protein torsinA, whose loss
of function causes the neurodevelopmental movement disorder DYT-TOR1A (DYT1) dystonia,
coordinates NPC spatial organization during this period without impacting total NPC density. Using a
new mouse line in which endogenous Nup107 is Halo-Tagged, we find that torsinA is essential for correct
localization of NPC formation. In the absence of torsinA, the inner nuclear membrane buds excessively
at sites of mislocalized, nascent NPCs, and NPC assembly completion is delayed. Our work implies that
NPC spatial organization and number are independently regulated and suggests that torsinA is critical
for the normal localization and assembly kinetics of NPCs.
INTRODUCTION
Nuclear pore complexes (NPCs) are large, evolutionarily
conserved structures that serve as the gateway between the
nucleus and cytoplasm, allowing passive diffusion of small
molecules and facilitating nucleocytosolic transport of
proteins and RNA1,2. Regulation of NPC biogenesis and
function is critical for coordinating information transfer
between the nucleus and cytoplasm, allowing cells to
dynamically respond to internal and external cues. Neurons
are heavily dependent on NPC function; neuronal plasticity
requires transport of signaling molecules and transcription
factors336 and mRNA export for local translation is essential
for several neurodevelopmental processes7311. Accordingly,
NPC defects are present in several nervous system
diseases12316 and mutations in nucleoporins cause early-
onset neurological illness17,18. Indeed, neurons face a
considerable challenge in maintaining proper nuclear
function throughout their lifetime. Whereas mitotic cells
disassemble and recreate the nucleus with each division,
neurons must manage nuclear pore number and localization
within a closed interphase nucleus. Neuronal NPCs exhibit
minimal turnover and nucleoporins (Nups) that constitute the
NPC are among the longest-lived proteins19,20, underscoring
the unique challenge neurons face in regulating NPC
formation and function. Despite the biological and clinical
importance of these events, little is understood about
neuronal NPC biogenesis. Furthermore, mechanisms
underlying NPC number, organization, and dynamics in both
neuronal and non-neuronal systems remain elusive.
TorsinA appears to lie at the intersection of neuronal
NPC biogenesis and neurodevelopmental disease. TorsinA
is a AAA+ protein that resides in the endoplasmic reticulum
(ER)/nuclear envelope (NE) lumen21329. The
neurodevelopmental movement disorder DYT-TOR1A
(DYT1) dystonia is caused by an in-frame 3-bp deletion in
the TOR1A gene that encodes a E-torsinA mutant
protein30,31. Several observations demonstrate that the NE is
an active site of torsinA activity. A <substrate trap= mutant of
torsinA primarily localizes to the NE24,25 and perturbing
torsinA levels causes changes in the LINC (linker of
nucleoskeleton and cytoskeleton) complex32,33.
Mislocalization of nuclear membrane proteins is also
observed in C. elegans germ cells lacking the torsin homolog
OOC-5, which additionally exhibit asymmetric plaques of
Nups34.
TorsinA-knockout (KO) or homozygous E mutant mice
develop abnormal NE evaginations or <blebs= exclusively in
post-migratory maturing neurons35, establishing E as a
loss-of-function (LOF) mutation. Biochemical studies are
consistent with a LOF effect of the E mutation36,37. NE blebs
are inner nuclear membrane (INM) outpouchings that project
into the NE lumen. These blebs occur transiently, emerging
.CC-BY-NC-ND 4.0 International licenseavailable under a
was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made
The copyright holder for this preprint (whichthis version posted April 27, 2023.;https://doi.org/10.1101/2023.04.26.538491doi:bioRxiv preprint
1
TorsinA is essential for the timing and localization of neuronal
nuclear pore complex biogenesis
Sumin Kim1,2, Sébastien Phan3, Thomas R. Shaw4, Mark H. Ellisman3, Sarah L. Veatch4, Sami J. Barmada1,2,*,
Samuel S. Pappas5,6,*, and William T. Dauer5,6,7,*
1Cellular and Molecular Biology Graduate Program, University of Michigan, Ann Arbor, MI; 2Department of Neurology, University of Michigan, Ann Arbor,
MI; 3National Center for Microscopy and Imaging Research, Center for Research on Biological Systems, Department of Neurosciences, School of
Medicine, University of California San Diego, La Jolla, CA; 4Department of Biophysics, University of Michigan, Ann Arbor, MI; 5Peter O9Donnell Jr. Brain
Institute, UT Southwestern, Dallas, TX; 6Department of Neurology, UT Southwestern, Dallas, TX; 7Department of Neuroscience, UT Southwestern,
Dallas, TX;
*To whom correspondence should be addressed: sbarmada@med.umich.edu; Samuel.Pappas@utsouthwestern.edu; William.Dauer@utsouthwestern.edu
Nuclear pore complexes (NPCs) regulate information transfer between the nucleus and cytoplasm. NPC
defects are linked to several neurological diseases, but the processes governing NPC biogenesis and
spatial organization are poorly understood. Here, we identify a temporal window of strongly upregulated
NPC biogenesis during neuronal maturation. We demonstrate that the AAA+ protein torsinA, whose loss
of function causes the neurodevelopmental movement disorder DYT-TOR1A (DYT1) dystonia,
coordinates NPC spatial organization during this period without impacting total NPC density. Using a
new mouse line in which endogenous Nup107 is Halo-Tagged, we find that torsinA is essential for correct
localization of NPC formation. In the absence of torsinA, the inner nuclear membrane buds excessively
at sites of mislocalized, nascent NPCs, and NPC assembly completion is delayed. Our work implies that
NPC spatial organization and number are independently regulated and suggests that torsinA is critical
for the normal localization and assembly kinetics of NPCs.
INTRODUCTION
Nuclear pore complexes (NPCs) are large, evolutionarily
conserved structures that serve as the gateway between the
nucleus and cytoplasm, allowing passive diffusion of small
molecules and facilitating nucleocytosolic transport of
proteins and RNA1,2. Regulation of NPC biogenesis and
function is critical for coordinating information transfer
between the nucleus and cytoplasm, allowing cells to
dynamically respond to internal and external cues. Neurons
are heavily dependent on NPC function; neuronal plasticity
requires transport of signaling molecules and transcription
factors336 and mRNA export for local translation is essential
for several neurodevelopmental processes7311. Accordingly,
NPC defects are present in several nervous system
diseases12316 and mutations in nucleoporins cause early-
onset neurological illness17,18. Indeed, neurons face a
considerable challenge in maintaining proper nuclear
function throughout their lifetime. Whereas mitotic cells
disassemble and recreate the nucleus with each division,
neurons must manage nuclear pore number and localization
within a closed interphase nucleus. Neuronal NPCs exhibit
minimal turnover and nucleoporins (Nups) that constitute the
NPC are among the longest-lived proteins19,20, underscoring
the unique challenge neurons face in regulating NPC
formation and function. Despite the biological and clinical
importance of these events, little is understood about
neuronal NPC biogenesis. Furthermore, mechanisms
underlying NPC number, organization, and dynamics in both
neuronal and non-neuronal systems remain elusive.
TorsinA appears to lie at the intersection of neuronal
NPC biogenesis and neurodevelopmental disease. TorsinA
is a AAA+ protein that resides in the endoplasmic reticulum
(ER)/nuclear envelope (NE) lumen21329. The
neurodevelopmental movement disorder DYT-TOR1A
(DYT1) dystonia is caused by an in-frame 3-bp deletion in
the TOR1A gene that encodes a E-torsinA mutant
protein30,31. Several observations demonstrate that the NE is
an active site of torsinA activity. A <substrate trap= mutant of
torsinA primarily localizes to the NE24,25 and perturbing
torsinA levels causes changes in the LINC (linker of
nucleoskeleton and cytoskeleton) complex32,33.
Mislocalization of nuclear membrane proteins is also
observed in C. elegans germ cells lacking the torsin homolog
OOC-5, which additionally exhibit asymmetric plaques of
Nups34.
TorsinA-knockout (KO) or homozygous E mutant mice
develop abnormal NE evaginations or <blebs= exclusively in
post-migratory maturing neurons35, establishing E as a
loss-of-function (LOF) mutation. Biochemical studies are
consistent with a LOF effect of the E mutation36,37. NE blebs
are inner nuclear membrane (INM) outpouchings that project
into the NE lumen. These blebs occur transiently, emerging
.CC-BY-NC-ND 4.0 International licenseavailable under a
was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made
The copyright holder for this preprint (whichthis version posted April 27, 2023.;https://doi.org/10.1101/2023.04.26.538491doi:bioRxiv preprint
100%
