A new human iPSC model of ALS: natural modifiers protecting FUS mutation carriers from the disease

肌萎缩性侧索硬化症（ALS）是一种复杂的神经退行性疾病

Amyotrophic lateral sclerosis (ALS) is a complex neurodegenerative disease with approximately 10- 15% familial cases caused by genetic mutations in an autosomal dominant fashion. Since familial and sporadic ALS are clinically indistinguishable, studies of familial ALS will facilitate understanding of ALS etiology in general. Among the ALS genes identified, several encode RNA binding proteins including Fused in Sarcoma (FUS). FUS functions in multiple RNA metabolic pathways. Mutant FUS protein is mis-localized to the cytoplasm where it forms granules and inclusions, a pathological hallmark of ALS. We and other groups have studied the FUS protein under physiological and pathological conditions in various models. Strikingly, we recently identified individuals in an extended ALS kindred who carry the ALS-linked FUS R521G mutation but live well beyond their 60s without developing ALS (Unaffected Mutation Carriers, UMCs). Our discovery of incomplete penetrance in this extended FUS-ALS pedigree is the first of its kind. We hypothesize that, despite carrying a disease-causing FUS mutation, UMCs have protective modifiers preventing disease development. The determination of such modifiers and the underlying mechanisms will point to novel therapeutic targets. Patient-derived iPSCs and their differentiated motor neurons (MNs) facilitate mechanistic studies in target cells in a relevant human genetic background. Taking advantage of newly generated iPSC lines derived from the unique ALS pedigree with multiple UMCs, we will characterize cellular and functional phenotypes of UMCs and determine the underlying protective molecular pathways. We propose two specific aims. We will define the cellular, biochemical and functional features protecting UMCs from developing ALS-like phenotypes using iPSC-differentiated MNs in Aim 1. We will differentiate MNs from iPSC lines of UMCs, ALS patients, and controls to characterize the pathophysiological dysfunctions at different differentiation stages using immunocytochemistry and electrophysiology. In addition, we will compare the cellular and functional features of iPSC-MNs from UMCs with ALS patients and controls, which will reveal which cellular and biochemical features are critical to protecting UMCs. In Aim 2, we will decipher biological pathways responsible for preserving normal functions in UMCs iPSC-MNs using RNA-Seq and whole genome sequencing analysis. The integration of phenotypic, transcriptomic and genomic data will provide in-depth understanding of how genetic modifiers function through molecular pathways to yield protective phenotypes. This MPI R21 project will be led by Dr. Ziyuan Guo at Cincinnati Children's Hospital Medical Center who has expertise on using stem cells and iPSCs as disease models and Dr. Haining Zhu at University of Kentucky who has expertise on ALS etiology. Completion of the proposed studies will provide novel insights into naturally occurring modifiers that protect UMCs from contracting ALS. The compensatory mechanism identified in this project will lay the foundation for future studies to develop future ALS therapies.