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Abstract

DNAJC13/RME-8 is a molecular co-chaperone which was originally identified as the mammalian homolog of receptor-mediated endocytosis 8 (rme-8) in a screen for endocytic defects in C. elegans (Y. Zhang, Grant, and Hirsh 2001). Subsequent investigation revealed a post-endocytic role for RME-8 in the sorting and trafficking of endosomal membrane. Mutations in the encoding gene, DNAJC13, have been linked to late-onset PD which is clinically indistinguishable from idiopathic PD (Vilariño-Güell et al. 2014). Similarly, multiple PD-associated genes encode proteins which functionally converge in endolysosomal sorting and trafficking pathways, including LRRK2, VPS35, and SNCA, which highlights the need for further investigation into this prominent pathomechanism (Farrer and Follett 2019). Thus, I have investigated disease-related phenotypes in the novel Dnajc13 p.N860S mutation (the equivalent of DNAJC13 p.N855S substitution in humans) knock-in mouse model (DKI). Initially, I describe an endosomal tubulation phenotype in primary cortical neuron cultures. This is consistent with previous observations in rme-8-deficient cells, suggesting Dnajc13 p.N860S leads to a loss-of-function that perturbs endosomal trafficking in DKI neurons. Next, in vivo analysis reveals alterations in the steady-state levels of the synaptic protein, α-Synuclein, and in the synaptic vesicle (SV) transporter, vesicular glutamate transporter 1 (Vglut1). To elaborate the mechanism responsible for these changes, I have identified a novel interaction between Dnajc13/Rme-8 and Clathrin Adaptor Protein complexes (AP-1, AP-2, and AP-3). Indeed, the interaction between mutant Dnajc13/Rme-8 and AP-2, a protein involved in Vglut1 recycling, is reduced in DKI neurons. As a corollary, I have observed an increased response to prolonged, repetitive stimulation in ex vivo field recordings, as well as changes in SV size and populations, which are consistent with altered α-Synuclein expression. Motor testing in young and aged animals reveals an age-associated motor phenotype in DKI animals. Compared to wild-type littermates, DKI mice exhibit worse motor performance and greater loss of dopaminergic soma in the substantia nigra, and increased pathologic spreading of toxic α-Synuclein aggregates when injected with α-Synuclein pre-formed fibrils. Together, these results provide insight into the underlying pathophysiology of the p.N855S mutation and support the designation of DNAJC13 as a PD gene, which has been an area of debate.