Isoform-level transcriptome Atlas of Macrophage Activation

Abstract

RNA-seq is routinely used to measure gene expression changes in response to cell perturbation. Genes that are up or down-regulated following perturbation in RNA-seq studies are designated as target genes for follow-up. However, RNA-seq is limited in its ability to capture the complexity of gene isoforms, defined by the exact composition of exons and transcription start sites (TSS) and poly(A) sites they contain, as well as the expression of these isoforms. Without knowing the composition of the most dominant isoform(s) of a target gene, a minority or non-existent isoform could be selected for follow-up solely based on available annotations for that target gene from databases that are incomplete, or by their nature not tissue specific, or do not provide key information on expression levels. In all, this can lead to loss in valuable resources and time. As the vast majority of genes in the human genome express more than one isoform, there is a great need to identify the complete range of isoforms present for each gene along with their corresponding levels of expression. Here, using the long-read nanopore-based R2C2 method, we generated an Isoform-level transcriptome Atlas of Macrophage Activation (IAMA) that identifies full-length isoforms in primary human monocyte-derived macrophages (MDMs). Macrophages are critical innate immune cells important for recognizing pathogens through use of Toll-like receptors (TLRs), culminating in the initiation of host defense pathways. We characterized isoforms for most moderate to highly expressed genes in resting and TLR-activated MDMs and generated a user-friendly portal built into the UCSC Genome Browser to explore the data (https://genome.ucsc.edu/s/vollmers/IAMA). Our atlas represents a valuable resource for innate immune research as it provides unprecedented isoform information for primary human macrophages.