Deciphering the In Vivo Huntingtin Interactome in the Mammalian Brain by Affinity Purification-Mass Spectrometry and Weighted Gene Correlation Network Analysis

摘要

Huntingtin (Htt), the protein mutated in Huntington's disease (HD), is a scaffold protein capable of interacting with hundreds of proteins, but its in vivo interactome in disease-relevant brain regions remains undefined. Using an affinity purification-mass spectrometry (AP-MS) based approach, we have performed a spatiotemporal proteomic profiling of in vivo full-length Htt interacting proteins in HD and wildtype mouse brains. We identified 749 Htt-interacting proteins that co-purified with Htt in distinct brain regions (cortex, striatum, or cerebellum) and age (2 or 12 months), and the interactome is significantly enriched with known Htt interactors. To gain a system-level view of these interactors, we performed Weighted Gene Correlation Network Analyses (WGCNA) of the entire proteomic dataset to identify protein correlation relationships among the Htt interactors. The analysis reveals an independently verifiable rank of proteins based on their correlation strength with Htt, and a robust network organization of distinct Htt-interacting protein modules in the brain. One interesting module (Red Module)containing Htt itself and 19 known interactors, is significantly enriched with genes implicated in neurological and psychological disorders beyond HD, and proteins involved in vesicular transport. We validated seven novel Red Module genes as Htt interactors in vitro and 13 genetic modifiers of HD in a Drosophila model. Together, our study demonstrates a novel WGCNA systems biology approach to decipher in vivo AP-MS proteomic datasets, and the value of such an approach to illuminate novel Huntington's disease protein networks in the brain.