Mechanisms of altered RNA transport dynamics by Kif1C mutations in heterospastic paraplegia

Emory University, Atlanta GA

Investigators

Abstract

Neuronal transport is essential for neuronal function and is often impaired in neurodegenerative diseases. We and others have identified impairments in mRNA transport and local translation in neurological diseases, including spinal muscular atrophy (SMA), Fragile X syndrome (FXS), amyotrophic lateral sclerosis (ALS) and most recently, myotonic dystrophy (DM1). Our recent research identified that the kinesin-3 motor, KIF1C, associates with MBNL1, an RNA binding protein (RBP) known to depleted in DM1. We characterized domains involved in MBN-KIF1C interactions involved in mRNA localization in neurons. This discovery has motivated the present research to explore a potential new link of RNA localization to heterospastic paraplegia type 58 (SPG58), a neurological disease caused by the aberrant function of KIF1C, although mechanisms are poorly understood. Our major goal is to test the hypothesis that KIF1C interacts with many RBPs to transport mRNAs in neurons and that these mechanisms are altered in SPG58. A critical gap is lack of a human neuronal disease model to investigate possible impairments in mRNA localization. We have recently developed human iPSC CRISPR induced KIF1C knockout cells that will be differentiated into motor neurons. We propose to use this neuronal disease model, together with a variety of imaging and biochemical methods, to identify a set of RBPs and their mRNA targets that depend on KIF1C. In Aim 1, we will identify RBP and mRNA cargoes specific to KIF1C. Aim 1A will test the hypothesis that KIF1C binds mRNAs through different RBP adaptors and domains and that this binding is altered by KIF1C SPG58 pathogenic variants. We will use mass spectrometry to broadly identify pathology-related RBP targets. Aim 1B will identify mRNAs mislocalized in neuronal processes of human neurons lacking KIF1C. We will bioinformatically compare the datasets obtained in Aim 1A and Aim 1B to identify specific KIF1C mRNP cargoes dysregulated in SPG58. In Aim 2, we will test the hypothesis that altered dynamics of KIF1C transport affects mRNA localization in neurons and are involved in HSP. In Aim 2A, we will perform live imaging of KIF1C, its pathogenic variants and known RBPs and mRNAs cargos in human motor neurons to evaluate KIF1C transport dynamics. In Aim 2B, we will likewise validate mRNA targets identified in Aim 1. Additionally, we will overexpress selected pathogenic Kif1C variants and evaluate rescue phenotypes of mRNP cargo localization. The proposed studies will uncover disease specific targets of KIF1C in a new human disease model and delve into the mechanism of dysregulated mRNA transport in SPG58 and its effect on neuronal function. This research will provide broader mechanistic insight into several neurological diseases with implications for development of therapeutic strategies.