High-content RNAi screening identifies the Type 1 inositol triphosphate receptor as a modifier of TDP-43 localization and neurotoxicity

Author： Kim Sang Hwa Zhan Lihong Hanson Keith A. Tibbetts Randal S.

Publisher： Oxford University Press

ISSN： 1460-2083

Source： Human Molecular Genetics, Vol.21, Iss.22, 2012-11, pp. : 4845-4856

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Abstract

Cytosolic aggregation of the nuclear RNA-binding protein (RBP) TDP-43 (43 kDa TAR DNA-binding domain protein) is a suspected direct or indirect cause of motor neuron deterioration in amyotrophic lateral sclerosis (ALS). In this study, we implemented a high-content, genome-wide RNAi screen to identify pathways controlling TDP-43 nucleocytoplasmic shuttling. We identified ∼60 genes whose silencing increased the cytosolic localization of TDP-43, including nuclear pore complex components and regulators of G2/M cell cycle transition. In addition, we identified the type 1 inositol-1,4,5-trisphosphate (IP3) receptor (ITPR1), an IP3-gated, endoplasmic reticulum (ER)-resident Ca²⁺ channel, as a strong modulator of TDP-43 nucleocytoplasmic shuttling. Knockdown or chemical inhibition of ITPR1 induced TDP-43 nuclear export in immortalized cells and primary neurons and strongly potentiated the recruitment of TDP-43 to Ubiquilin-positive autophagosomes, suggesting that diminished ITPR1 function leads to autophagosomal clearance of TDP-43. The functional significance of the TDP-43-ITPR1 genetic interaction was tested in Drosophila, where mutant alleles of ITPR1 were found to significantly extended lifespan and mobility of flies expressing TDP-43 under a motor neuron driver. These combined findings implicate IP3-gated Ca²⁺ as a key regulator of TDP-43 nucleoplasmic shuttling and proteostasis and suggest pharmacologic inhibition of ITPR1 as a strategy to combat TDP-43-induced neurodegeneration in vivo.