Proteomic characterization of the Alzheimer’s disease risk factor BIN1 interactome

ABSTRACT: The gene BIN1 is the second-largest genetic risk factor for late-onset Alzheimer’s disease (LOAD). BIN1 is expressed in neurons and glia in the brain as multiple isoforms, including neuron-specific and ubiquitously expressed isoforms. BIN1 is an adaptor protein that regulates membrane dynamics in many cell types. Previously, we reported that BIN1 predominantly localizes to presynaptic terminals in neurons and regulates presynaptic vesicular release. However, the function of neuronal BIN1 as it relates to LOAD is not fully understood. A fundamental gap in the field is an unbiased characterization of neuronal BIN1-interacting proteins and proximal neighbors. To fill this void and help define neuronal BIN1’s functions in the brain, we applied TurboID-based proximity labeling to identify proteins biotinylated by neuronal BIN1 isoform 1-TurboID fusion protein (BIN1iso1-TID) in cultured mouse neuroblastoma (N2a) cells in vitro and in adult mouse brain neurons in vivo. Label-free proteomics identification of BIN1iso1-TID biotinylated proteins resulted in the discovery of 361 proteins from the N2a cells and 897 proteins in mouse brain neurons as BIN1iso1-associated (proximal) or interacting proteins. A total of 92 proteins were common in the two datasets, indicative that these are high-confidence BIN1-interacting or proximity proteins. SynapticGO analysis of the mouse brain dataset showed that BIN1iso1-TurboID labeled 159 synaptic proteins, with 60 corresponding to the synaptic vesicle cycle. Based on a phosphopeptide analysis of the neuronal BIN1iso1-TID interactome and kinase prediction, we chose to validate AAK1, CDK16, SYNJ1, PP2BA, and RANG through immunostaining and proximity ligation assays as members of the BIN1 interactome in the mouse brain. By identifying a number of previously unknown BIN1 proximal and potential interacting proteins, this study lays a foundation for further investigations on neuronal BIN1 function.

INSTRUMENT(S):

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Brain, Cell Culture

SUBMITTER: Stanley Stevens  

LAB HEAD: Dr. Stanley Stevens, Jr.

PROVIDER: PXD060638 | Pride | 2025-08-22

REPOSITORIES: Pride