Research Project
10240686
Summary The Human Correlation Core will determine whether the novel genes identified by Projects 1 and 2 of this U19 are relevant to human disease. During the preceding fifteen years, the investigators forming the U19 team have identified and contextualized a large number of previously unknown immune regulatory genes, both by forward genetics studies and/or systems biology approaches. In addition, systems approaches have been used to place novel genes within immune regulatory networks. These have included the molecular pathways involved in pathogen recognition, signal transduction, and gene regulation leading to appropriately tailored responses to infection. Over the years, this program has provided seminal information about multiple pathways that mediate inflammation, regulate responses to infection, and underlie successful vaccination. Here the Core will test these genes for relevance in human immunity. The efforts of the Human Correlation Core will be greatly expanded by systems based network analysis (Project 2) and molecular phenotyping (Signaling Core). The Aderem laboratory (Project 2) has established several methods for global transcriptomic analysis of limiting samples including robust amplification protocols for handling small quantities of mRNA and chromatin accessibility measures (ATAC-seq) that can interrogate as few as 500 cells. During the previous funding period of this U19 program, the Signaling Core developed single-cell mass cytometry (CyTOF) as well as methods to both efficiently implement this technology at high- throughput and comprehensively analyze the resulting data. CyTOF permits the simultaneous measurement of 50-100 parameters including surface markers, cytokines and chemokines, and phosphorylated signaling molecules at the single-cell level. Where appropriate, the Core will collaborate with the Aderem and Nolan laboratories to apply these approaches to primary human cells. Since some of our technologies, for example CRISPR/Cas9 gene editing, require large numbers of cells as starting material (it is particularly difficult to use the technique in non-dividing myeloid cells) the Core has established new approaches to isolate large numbers of macrophage lineage cells derived from CD34+ cord blood cells.
