Abstract
Most cases of early onset torsion dystonia (DYT1) are caused by a 3-base pair deletion in one allele of the TOR1A gene causing loss of a glutamate in torsinA, a luminal protein in the nuclear envelope. This dominantly inherited neurologic disease has reduced penetrance and no other medical manifestations. It has been challenging to understand the neuronal abnormalities as cells and mouse models which are heterozygous (Het) for the mutant allele are quite similar to wild-type (WT) controls. Here we found that patient fibroblasts and mouse neurons Het for this mutation showed significant differences from WT cells in several parameters revealed by infection with herpes simplex virus type 1 (HSV) which replicates in the nucleus and egresses out through the nuclear envelope. Using a red fluorescent protein capsid to monitor HSV infection, patient fibroblasts showed decreased viral plaque formation as compared to controls. Mouse Het neurons had a decrease in cytoplasmic, but not nuclear HSV fluorescence, and reduced numbers of capsids entering axons as compared to infected WT neurons. These findings point to altered dynamics of the nuclear envelope in cells with the patient genotype, which can provide assays to screen for therapeutic agents that can normalize these cells.
Highlights
In order to develop new therapies for dystonia it is important to generate assays to screen for drugs or genes that can normalize DYT1 genotypic cells
We found a decrease in viral plaque number and size in DYT1 compared to control fibroblasts, and decreased replication of Herpes Simplex Virus type 1 (HSV) in neurons homozygous for the DYT1 mutation (KI) compared to Het or WT
Based on the findings of others that cells with alterations in torsinA expression showed decreased ability of HSV to replicate, and to exit the NE15–18, we evaluated this for the first time in skin fibroblast cultures derived from DYT1 patients and controls
Summary
In order to develop new therapies for dystonia it is important to generate assays to screen for drugs or genes that can normalize DYT1 genotypic cells. We took advantage of a replication competent variant of HSV in which a capsid protein, VP26, is fused to monomeric red fluorescent protein (RFP-VP26)[29] This variant HSV was used to monitor plaque number and size in human DYT1 and control fibroblasts. We found a decrease in viral plaque number and size in DYT1 compared to control fibroblasts, and decreased replication of HSV in neurons homozygous for the DYT1 mutation (KI) compared to Het or WT. Both Het and KI neurons showed a decrease in nuclear egress of the HSV capsids into the cytoplasm, as compared to WT neurons. HSV provides a probe to distinguish the WT and Het genotypes, with the latter being genotypically similar to DYT1 patients, with significant parameters including reduced propagation in patient fibroblasts, and reduced viral fluorescence in the cell body in Het, as compared to WT neurons
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