Electronic Thesis and Dissertation Repository

Degree

Master of Science

Program

Pathology

Supervisor

Michael J. Strong

Abstract

Rho Guanine Nucleotide Exchange factor (RGNEF) is a 180 kDa protein which forms pathological neuronal cytoplasmic inclusions in degenerating spinal motor neurons in amyotrophic lateral sclerosis (ALS) and for which the gene (ARHGEF28) is mutated in a subset of cases. Despite having previously been shown to localize to the nucleus and to undergo a nuclear cytosolic shift in response to cellular injury, the mechanism of its nuclear import has yet to be elucidated. Here we use site-directed mutagenesis with a combination of subcellular fractionation and confocal microscopy to identify a functional nuclear localization signal (NLS) within the Pleckstrin Homology (PH) domain of RGNEF. We show that the function of the NLS is conserved both in full length RGNEF and when expressed as only the PH domain. This advances the growing body of literature implicating nuclear mislocalization of ALS-related proteins as a feature of ALS pathology. Additionally, this is the first PH domain-embedded NLS identified, despite the prevalence of PH domains in the human proteome and the similarities in their consensus motifs.

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