Upregulation of RIN3 induces endosomal dysfunction in Alzheimer’s disease

Abstract

Background In Alzheimer’s Disease (AD), about one-third of the risk genes identified by GWAS encode proteins that function predominantly in the endocytic pathways. Among them, the Ras and Rab Interactor 3(RIN3) is a guanine nucleotide exchange factor (GEF) for the Rab5 small GTPase family and has been implicated to be a risk factor for both late onset AD (LOAD) and sporadic early onset AD (sEOAD). However, how RIN3 is linked to AD pathogenesis is currently undefined. Methods Quantitative PCR and immunoblotting were used to measure the RIN3 expression level in mouse brain tissues and cultured basal forebrain cholinergic neuron (BFCNs). Immunostaining was used to define subcellular localization of RIN3 and to visualize endosomal changes in cultured primary BFCNs and PC12 cells. Recombinant flag-tagged RIN3 protein was purified from HEK293T cells and was used to define RIN3-interactomes by mass spectrometry. RIN3-interacting partners were validated by co-immunoprecipitation, immunofluorescence and yeast two hybrid assays. Live imaging of primary neurons was used to examine axonal transport of amyloid precursor protein (APP) and β-secretase 1 (BACE1). Immunoblotting was used to detect protein expression, processing of APP and phosphorylated forms of Tau. Results We have shown that RIN3 mRNA level was significantly increased in the hippocampus and cortex of APP/PS1 mouse brain. Basal forebrain cholinergic neurons (BFCNs) cultured from E18 APP/PS1 mouse embryos also showed increased RIN3 expression accompanied by early endosome enlargement. In addition, via its proline rich domain, RIN3 recruited BIN1(bridging integrator 1) and CD2AP (CD2 associated protein), two other AD risk factors, to early endosomes. Interestingly, overexpression of RIN3 or CD2AP promoted APP cleavage to increase its carboxyl terminal fragments (CTFs) in PC12 cells. Upregulation of RIN3 or the neuronal isoform of BIN1 increased phosphorylated Tau level. Therefore, upregulation of RIN3 expression promoted accumulation of APP CTFs and increased phosphorylated Tau. These effects by RIN3 was rescued by the expression of a dominant negative Rab5 (Rab5S34N) construct. Our study has thus pointed to that RIN3 acts through Rab5 to impact endosomal trafficking and signaling. Conclusion RIN3 is significantly upregulated and correlated with endosomal dysfunction in APP/PS1 mouse. Through interacting with BIN1 and CD2AP, increased RIN3 expression alters axonal trafficking and procession of APP. Together with our previous studies, our current work has thus provided important insights into the role of RIN3 in regulating endosomal signaling and trafficking.

Identification of DNA Methylation Changes That Predict Onset of Post-traumatic Stress Disorder and Depression Following Physical Trauma

“…We also report that Ras and Rab interactor 3 (RIN3), receptor-type tyrosine-protein phosphatase 2 (PTPRN2), and long intergenic non-protein coding RNA 319 (LINC00319) genes from our prior analyses with MeDIP-seq on PTSD patients were also present as significant genes in this analysis (Martin et al, 2018). The association of these genes and neurodegenerative disorders have been reported previously (Boden et al, 2017;Shen et al, 2020). In particular, RIN3 is a guanidine nucleotide exchange factor have been significantly associated with Alzheimer's disease (AD) pathology via endosomal dysfunction (Shen et al, 2020).…”

A novel age-informed approach for genetic association analysis in Alzheimer’s disease

“…Common variants in a locus near RIN3 and SLC24A4 , were reported to be associated with AD susceptibility [2]. Increased RIN3 expression in APP/PS1 mouse models was shown to correlate with endosomal dysfunction and altered axonal trafficking and processing of APP [60]. For these reasons, the Rab related proteins involved in the endolysosomal and retromer pathways have been considered as promising therapeutic targets for AD [54].…”

Exome sequencing identifies rare damaging variants in ATP8B4 and ABCA1 as novel risk factors for Alzheimer’s Disease

“…SRC has been described to potentially modulate APP trafficking/metabolism 12 , but also Tau phosphorylation 13 . RIN3 has been associated with endosomal dysfunction and APP trafficking/metabolism 14,15 . CLU (also known as APOJ) has been found to affect Aβ aggregation and clearance 16 and ACE is suggested to have a role in Aβ degradation 17 .…”