1. Determine the molecular mechanisms of repeat RNA metabolisms and RAN translation
Nucleotide repeat expansions underlie a heterogeneous groups of neurological and neuromuscular disorders with various disease mechanisms depending on the gene function, structure and location of the expansion. Hexanucleotide repeat expansion in the C9orf72 gene is the most frequent inherited cause of both ALS and FTD. A leading hypothesis for disease mechanism is gain of toxicity from the expanded repeats, which are transcribed in both sense and antisense directions and give rise to distinct sets of intranuclear RNA foci. This could sequester certain RNA-binding proteins and lead to their loss of function. An alternative but not mutually exclusive mechanism is the aberrant accumulation of dipeptide repeat proteins produced by repeat-associated non-ATG (RAN) translation in all six reading frames (poly-GA, poly-GR, poly-PA, poly-PR and poly-PG) of both strands. A third hypothesis is the expansions cause gene silencing and lead to haploid insufficiency of C9orf72 protein. Till today, there is no established evidence supporting what is the main factor driving the disease pathogenesis. Our research goal is to use biochemical, proteomic, genomic and screening approaches to decipher the various mechanisms of repeat expansion mediated toxicity and advance therapeutic target development, especially by applying RNA Biology knowledge and technologies.
2. Decipher the global RNA metabolism dysregulation in ALS/FTD
Many disease causative genes in ALS and FTD are linked to RNA regulation, including C9ORF72 repeat expansion and RNA-binding proteins (RBPs), such as TDP-43 and FUS/TLS. Besides causative mutations found in familial ALS, the pathology of RBPs is widely found in sporadic ALS and FTD. Understanding how the endogenous RNA metabolism pathways are affected by the mutated or pathological RBPs and expanded repeats will likely provide novel insights on biomarker and therapeutic targets development. Neurons have extremely complex RNA metabolism and highly specialized RNA processing pathways, probably due to their highly complex morphologies and functions. We are interested in deciphering the global RNA metabolism dysregulation by high-throughput sequencing techniques, including splicing, translation, modification, and the connection to epigenetic regulation via ncRNAs.
3. Identify disease-modifying genes by CRISPR screening in human iPSC-derived neurons
Genetic screenings carried out on model organisms such as yeast, flies and nematodes have played important roles in understanding gene functions and the pathogenic mechanisms induced by disease-causative mutations, which facilitate the development of novel therapeutic targets. We have established the cutting-edge CRISPR-Cas9 screening platform to screen for modifiers directly in human cells and iPSC-derived neurons. We develop assays to screen for modifiers of specific molecular pathways (RNA processing, protein homeostasis), as well as ones that can inhibit mutant gene-induced toxicity and improve neuron survival. Identification of druggable gene products will facilitate treatment development of neurodegenerative diseases.
