Defining new targets in lupus through identification of non-coding SNPs
Scientists have found more than 50 regions of DNA that contribute to lupus risk. At least 35 of these DNA regions regulate gene activity—they serve as magnets that attract proteins, which in turn, act as control switches that determine whether a gene is more or less active. Variations in gene activity influence a person’s biology—for example, whether they are more or less likely to develop a disease like lupus. The TIL grant will enable Dr. Nigrovic to study each of these 35 DNA control centers to determine how they contribute to lupus risk.
Dr. Nigrovic will identify the proteins that interact with the DNA regions using state-of-the-art technologies. Once the control proteins are known, he will be able to build up a more complete picture of the genes and biological pathways that cause lupus.
What this study means to people with lupus
This line of research is expected to discover new targets for treatments to prevent or cure lupus.
Complex diseases such as systemic lupus erythematosus arise through the interplay of many different genes. Genome-wide association studies (GWAS) have defined more than 50 specific loci that carry lupus risk. Each of these “hits” marks a pathway that plays a causative role in lupus, and therefore represents a potential therapeutic target. Unfortunately, linked haplotypes are typically large and rarely contain coding variants, rendering the identification of causative genetic polymorphisms unexpectedly difficult, since most appear to be regulatory. Finding these regulatory variants, and then identifying proteins such as transcription factors that mediate their action, has proven to be a daunting challenge.
To address this challenge, our lab has developed two new molecular techniques that enable us to bridge the gap between genetic data and disease biology. The first, termed SNP-seq, begins with the generation of DNA constructs for each potentially regulatory single nucleotide polymorphism (SNP). Thousands of such individual constructs are then incubated with nuclear proteins, followed by use of enzymatic restriction to identify those that are sheltered by binding proteins. Following molecular confirmation, we then employ Flanking Restriction Enhanced Pulldown (FREP, PLOS Genetics 2016) to identify specific binding proteins using mass spectrometry. Together, SNP-seq and FREP provide a new pathway from genetic data to mechanism.
Here we propose to use these methods to identify new therapeutic targets in lupus. Our analysis of GWAS data has identified 35 loci that are likely to contain regulatory variants. In Aim I, we will apply SNP-seq to these regions, and use state-of-the-art bioinformatics to narrow down a set of specific targets that we will then validate experimentally. In Aim II, we will use FREP to define specific proteins that bind functional variants that we have already identified, and then more broadly to identify proteins and pathways implicated by our lupus-wide genetic analysis. Using this approach, we aim to expose a large range of previously unexpected biological pathways that cause human lupus, thereby opening up new frontiers in disease treatment.