Research
RNA folding of a 69 bp repeat in WDR7
Gene discovery and gene therapy for Amyotrophic Lateral Sclerosis
The burden of neurodegenerative diseases such as Amyotrophic Lateral Sclerosis (ALS) is becoming increasingly apparent as our global population ages. Two-thirds of individuals with familial ALS have a causative mutation in one of several genes including SOD1 (10-20%) TARDBP and FUS (1-5%) or hexanucleotide repeat expansions in C9orf72 (~40%). We were one of the first two groups to identify that mutations in TARDBP are causative in a subset of patients with ALS and that the mutations lead to accumulation of C-terminal truncated fragments of the TDP-43 protein. We are working towards identifying genetic contributions that cause or modify susceptibility to ALS including a 69-bp tandem repeat expansion in an intron of WDR7.
Research projects of interest:
Novel genetic contributions to ALS
We are using a combination of approaches including long-read sequencing and whole genome sequence analysis to identify novel contributors to disease, with a particular focus on tandem repeat expansions. Identifying the contributions of tandem repeats can help explain some of the missing heritability in individuals with ALS. Understanding the role of repeat expansions can reveal mechanisms associated with ALS and provide new therapeutic avenues for discovery.
2. Approaches for neuronal gene therapy
The genetics for a substantial subset of cases in ALS is firmly established meaning that a gene-specific tailored therapeutic approach has the potential to be utilized. We are interested in leveraging our experience in delivering shRNAs to determine the ability to suppress genes that are causative for neurodegenerative disease in the brain, and the consequence on existing neuronal microRNAs.
Non-coding RNA biology in Alzheimer's disease (AD)
An early hallmark of Alzheimer’s disease is loss of neuronal synapses. To better comprehend the transcriptional mechanisms associated with synapse loss, we perform RNA sequencing of synapses in the form of synaptosomes in individuals with AD compared to neurologically normal individuals. We find differentially expressed mRNAs and widespread changes in noncoding RNAs – including circular RNAs – in AD synapses.
Alzheimer's disease genetics projects
We are interested in several other questions pertaining to AD biology including the role of cryptic exon splicing events in sporadic AD (Course et al., Brain, 2023) and the role of tandem repeat expansions as AD genetic risk factors.
Safe and effective small hairpin RNA delivery for liver gene therapy
RNA interference is a powerful tool to disrupt virtually any gene of interest. RNA interference and other gene therapy approaches represent promising tools to treat various disorders. However, parameters need to be established to ensure safe and effective gene knockdown. Recombinant adeno-associated virus vectors (rAAVs) are at the forefront in the long road towards regulatory approval for gene therapy. They are safe, and AAVs typically do not integrate into the genome but persist in episomal state and can transduce the vast majority of hepatocytes.
We are working on devising mechanisms to safely deliver robust small hairpin RNAs (shRNAs) while sustaining gene knockdown by RNA interference. By high-throughput sequencing of delivered shRNAs and existing microRNAs, we have identified a signature of high shRNA expression and the consequence of one microRNA, miR-122, in particular. The effect on miR-122 levels, notably the various abundant isoforms of miR-122-5p, has established guidelines for identifying and interpreting other situations where miR-122 isoforms are altered.
Imprinted microRNA clusters that are aberrantly activated in liver cancer and in the absence of miR-122
MicroRNAs are small RNA species that mediate site-specific repression of mRNA targets. The number of identified microRNAs is about an order of magnitude less than the number of genes and their corresponding mRNAs. Furthermore, while each mRNA species constitutes a small fraction of the total mRNA transcripts in a cell, certain microRNAs have a much greater representation of the total microRNA pool and can have a more drastic effect on their cellular microRNA counterparts. This is especially notable in the liver where one major microRNA, miR-122, accounts for the bulk (~70%) of liver microRNA read counts yet estimates of the most abundant mRNAs account for only a few percent of total mRNA abundance.
We have identified a cluster of ~55 microRNAs at the Dlk1-Dio3 locus on mouse chromosome 12qF1 that are induced in Kras-induced lung adenocarcinoma (highlighted in red in image to the right). This same locus is activated in hepatocellular carcinoma, particularly due to AAV virus integration events. Through CRISPR/Cas9 approaches we are interested in how the cluster of microRNAs becomes aberrantly regulated.
Questions we are particularly interested in addressing are the following: is one or more microRNA specifically responsible for oncogenesis ? What is the mechanism by which AAV integrates and activates the locus? Are there ways to inactivate this locus in the event the region becomes aberrantly expressed?
