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O1‐05‐02: Role of PICALM in APP internalization, Abeta production and plaque pathogenesis
Author(s) -
Cirrito John,
Xiao Qingli,
Lee JinMoo,
Wang Yan,
Gil So Chon,
Yan Ping,
Gonzales Ernie,
Perez Ron
Publication year - 2011
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2011.05.257
Subject(s) - gene knockdown , microbiology and biotechnology , internalization , biology , small hairpin rna , cancer research , cell culture , cell , biochemistry , genetics
Background: At ICAD 2009, much excitement was generated by the announcement of several genes that were associated with late onset Alzheimer’s disease (AD) in the first round of genome-wide association studies. The association of PICALM (phosphatidylinositol clathrin assembly lymphoidmyeloid leukemia) with AD was novel and unexpected. In this study, we sought to determine the role of PICALM inAD pathogenesis.Methods:Because PICALM gene deletion results in embryonic lethality due to brain maldevelopment, we took a gene knockdown approach using shRNA. Knockdown and overexpression of PICALMwere performed in Neuroblastoma N2a-APP695 cells. APP internalization was measured after labeling cell surface proteins with biotin. Western blotting was used to detect internalized APP, CTFa and CTFb. Extracellular Aßx-42 levels were measured with ELISA from conditioned medium. In vivo, PICALM expression was altered using AAV8 viral gene transfer of shRNA or PICALM-HA into the hippocampus of 6 month old APP/PS1 mice. Aß concentration and amyloid plaque load were measured 4 months later. Results: PICALM colocalized with APP in intracellular vesicles of N2a-APP695 cells only after endocytosis was initiated. PICALMknockdown in N2a-APP695 cells resulted in reduced APP internalization and Aß42 generation, as measured in conditioned medium (1302 pg/ml6 18.2 vs 1003 pg/ml6 39.1, p< 0.005). Conversely, PICALM overexpression increased APP internalization and Aß42 production (1124 pg/ml 6 28.6 vs 1388 pg/m l 6 54.6, p 1⁄4 0.01). In vivo, viral gene transfer of PICALM shRNA into the hippocampus of APP/PS1 mice decreased PBS-soluble Aß42 levels (363 pg/ml 6 44 vs 253 pg/ml 6 17, p < 0.05) and amyloid plaque load (1.11% 6 0.09 vs 0.79% 6 0.12, p < 0.05). Conversely, overexpression of PICALM-HA increased Aß42 levels (317 pg/ml6 49 vs 499 pg/ml6 68, p< 0.05) and amyloid plaque load (1.15% 6 0.07 vs 1.53% 6 0.07, p < 0.005). Conclusions: PICALM, an adaptor protein involved in clathrin-mediated endocytosis, regulates APP internalization and subsequent Aß generation. Furthermore, PICALM plays a role in amyloid plaque pathogenesis and may provide a novel therapeutic target for AD.