Recently, the localization of amyloid precursor protein (APP) into reversible nanoscale supramolecular assembly or “nanodomains” has been highlighted as crucial towards understanding the onset of the molecular pathology of Alzheimer’s disease (AD). Surface expression of APP is regulated by proteins interacting with it, controlling its retention and lateral trafficking on the synaptic membrane. Here, we evaluated the involvement of a key risk factor for AD, PICALM, as a critical regulator of nanoscale dynamics of APP. Although it was enriched in the postsynaptic density, PICALM was also localized to the presynaptic active zone and the endocytic zone. PICALM colocalized with APP and formed nanodomains with distinct morphological properties in different subsynaptic regions. Next, we evaluated if this localization to subsynaptic compartments was regulated by the C-terminal sequences of APP, namely, the “Y682ENPTY687” domain. Towards this, we found that deletion of C-terminal regions of APP with partial or complete deletion of Y682ENPTY687, namely, APP–Δ9 and APP–Δ14, affected the lateral diffusion and nanoscale segregation of APP. Lateral diffusion of APP mutant APP–Δ14 sequence mimicked that of a detrimental Swedish mutant of APP, namely, APP–SWE, while APP–Δ9 diffused similar to wild-type APP. Interestingly, elevated expression of PICALM differentially altered the lateral diffusion of the APP C-terminal deletion mutants. These observations confirm that the C-terminal sequence of APP regulates its lateral diffusion and the formation of reversible nanoscale domains. Thus, when combined with autosomal dominant mutations, it generates distinct molecular patterns leading to onset of Alzheimer’s disease (AD).
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The authors thank the imaging facility at the Division of Biological Sciences, IISc Bengaluru, India and IISER Pune, India. We also thank the Central Animal Facility at IISc, Bangalore, India. We thank Sandhya Subramaniam for her help with primary neuronal cultures. We thank Aishwarya Babu for contribution to protein purification and biochemical analysis. We thank Dr. Narendrakumar Ramanan for sharing resources and help with molecular strategies and technical assistance. We thank Dr. Jean-Baptiste Sibarita for helpful discussion and for sharing advanced analysis modules. We also thank Dr. David Perrais for his insightful comments on the functional molecular markers at endocytic zone.
This work was supported by generous grants from Department of Biotechnology (Innovative Biotechnologist Award to D.N., M.J.), Department of Biotechnology Genomics Engineering Taskforce (D.N.), Ramalingaswami Fellowship to D.N. and M.J.), DBT-IISc Partnership program (to D.N.), IISc-STAR program grant to D.N., CBR-FABRIC GRANT (DN), Science and Engineering Research Board (SERB-ECR award to M.J. CRG CRG/2019/006899 and CRG/2022/002726 to D.N.), Indian Institute of Science (Institute of Excellence Program) and University Grants Commission, India grants to D.N. and Tata Trusts, India program grant to D.N (Co-investigator). We also acknowledge and thank; DBT (BT/PR32489/BRB/10/1786/2019) and SERB (CRG/2019/000281) for S.R.G.S. V.B. and M.S.H.S thank UGC for JRF and SRF fellowships. S.K. thanks postdoctoral fellowship support through Department of Biotechnology, Genomics Engineering Taskforce grant awarded to D.N and N.S. thanks senior research fellowship from CSIR, India.
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Wild type Sprague–Dawley Pups were acquired from Central Animal Facility (CAF, IISc). The pups from postnatal day-0 (P0), were sacrificed to dissect the hippocampus for primary neuronal cultures. All the procedures in this study were performed according to the rules and guidelines declared in the Compendium of CPCSEA 2018 by the Committee for the Purpose of Control and Supervision of Experimental Animals (CPCSEA), Ministry of Fisheries, Animal Husbandry and Dairying, India. The research protocol (CAF/Ethics/659 and CAF/Ethics/790) was approved by the Institutional Animal Ethics Committee (IAEC) of the Indian Institute of Science.
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Belapurkar, V., Mahadeva Swamy, H.S., Singh, N. et al. Real-time heterogeneity of supramolecular assembly of amyloid precursor protein is modulated by an endocytic risk factor PICALM. Cell. Mol. Life Sci. 80, 295 (2023). https://doi.org/10.1007/s00018-023-04939-w