Early-onset dystonia is associated with the deletion of one of a pair of glutamic acid residues (c.904_906delGAG/c.907_909delGAG; p.Glu302del/Glu303del; ΔE 302/303) near the carboxyl-terminus of torsinA, a member of the AAA+ protein family that localizes to the endoplasmic reticulum (ER) lumen and nuclear envelope (NE). This deletion commonly underlies early-onset DYT1 dystonia. While the role of the disease-causing mutation, torsinAΔE, has been established through genetic association studies, it is much less clear whether other rare human variants of torsinA are pathogenic. Two missense variations have been described in single patients; R288Q (c.863G>A; p.Arg288Gln; R288Q) identified in a patient with onset of severe generalized dystonia and myoclonus since infancy, and F205I (c.613T>A, p.Phe205Ile; F205I) in a psychiatric patient with late-onset focal dystonia. In this study, we have undertaken a series of analyses comparing the biochemical and cellular effects of these rare variants to torsinAΔE and wild-type (wt) torsinA in order to reveal whether there are common dysfunctional features. The results revealed that the variants, R288Q and F205I, are more similar in their properties to torsinAΔE protein than to torsinAwt. These findings provide functional evidence for the potential pathogenic nature of these rare sequence variants in the TOR1A gene, thus implicating these pathologies in the development of dystonia.
Background
Microfluidic platforms for quantitative evaluation of cell biologic
processes allow low cost and time efficient research studies of biological
and pathological events, such as monitoring cell migration by real-time
imaging. In healthy and disease states, cell migration is crucial in
development and wound healing, as well as to maintain the body's
homeostasis.
New Method
The microfluidic chambers allow precise measurements to investigate
whether fibroblasts carrying a mutation in the TOR1A gene,
underlying the hereditary neurologic disease - DYT1 dystonia, have decreased
migration properties when compared to control cells.
Results
We observed that fibroblasts from DYT1 patients showed abnormalities
in basic features of cell migration, such as reduced velocity and
persistence of movement.
Comparison with Existing Method
The microfluidic method enabled us to demonstrate reduced
polarization of the nucleus and abnormal orientation of nuclei and Golgi
inside the moving DYT1 patient cells compared to control cells, as well as
vectorial movement of single cells.
Conclusion
We report here different assays useful in determining various
parameters of cell migration in DYT1 patient cells as a consequence of the
TOR1A gene mutation, including a microfluidic platform,
which provides a means to evaluate real-time vectorial movement with single
cell resolution in a three-dimensional environment.
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