Young, Jessica EKnupp, Allison2022-01-262022-01-262022-01-262021Knupp_washington_0250E_23634.pdfhttp://hdl.handle.net/1773/48165Thesis (Ph.D.)--University of Washington, 2021Alzheimer’s disease (AD) is a progressive neurodegenerative disorder, and the most common cause of dementia among adults. There is currently no treatment that halts disease progression. There is ample pathological and biological evidence for endolysosomal network (ELN) dysfunction in AD, and emerging genetic studies repeatedly implicate ELN genes such as SORL1 as associated with increased AD risk. The SORL1 gene encodes the protein SORLA, a sorting receptor involved in retromer-related endosomal traffic. Many SORL1 genetic variants increase AD risk, and rare loss-of-function truncation mutations have been found to be causal of AD. To model the causal loss-of-function mutations, we used CRISPR/Cas9 technology to deplete SORL1 in human induced pluripotent stem cells (hiPSCs) to test the hypothesis that loss of SORL1 (SORL1 KO) contributes to AD pathogenesis by leading to dysfunction in ELN trafficking. We additionally used CRISPR/Cas9 to insert AD-risk variants in the VPS10 domain of SORL1 (SORL1Var) in hiPSCs to test the hypothesis that these VPS10 variants result in loss of SORLA function and lead to ELN dysfunction. We report that loss of SORL1 as well as SORL1 VPS10 variants in hiPSC-derived neurons leads to early endosome enlargement, a cellular phenotype that is indicative of ‘traffic jams’ and is now considered a hallmark cytopathology AD. We further report trafficking defects in the recycling and degradative pathways of the ELN in SORL1 KO neurons. Finally, we determine that retromer stabilizing small molecules reduce early endosome enlargement in both SORL1 KO and SORL1Var neurons. Collectively, and together with other recent observations, these findings suggest that SORL1 is a key and broad regulator of ELN trafficking in neurons, a conclusion that has both pathogenic and therapeutic implications. Moreover, demonstrating a partial rescue of cellular phenotypes in SORL1 deficient neurons will contribute to the development of new and precision treatments for AD.application/pdfen-USCC BYPathologyMolecular medicineThesis