Hierarchical Control in an Endocrine-mediated Gene Regulatory Network Supporting Innate Immunity

About This Grant

PROJECT SUMMARY Hormones pleiotropically regulate diverse physiological processes, often through tiered gene regulatory networks (GRNs) which consist of primary response genes that are directly regulated by the hormone and secondary response genes that are regulated by transcription factors in the primary response set. A central hypothesis in this proposal is that pleiotropic hormones can enact distinct gene regulatory programs via hierarchically tiered gene regulation. It is further hypothesized that modest differences in the primary level of regulation can result in larger differences at the secondary level, leading to functionally specialized GRNs over development and in response to environmental stimulus. However, distinguishing primary from secondary regulation is difficult or impossible from transcriptomic data alone, presenting a major hindrance to deciphering how hormones regulate pleiotropic balances. This project will overcome that challenge by combining transcriptomic profiling with computational analysis and direct assays of DNA-binding to decipher primary and secondary levels of gene regulation. The project will focus on the hormone 20- hydroxyecdysone (or ecdysone, 20E) in the Drosophila melanogaster model system. 20E has well-defined roles in development and a documented but undefined role in regulating the immune system. In Aim 1, an ex vivo tissue assay will be used to build the 20E gene-regulatory network. Transcriptomic analysis, measurement of direct DNA binding by the nuclear hormone receptor, and characterization of the chromatin landscape will be combined to identify primary targets of 20E signaling. Transcription factors in the primary-regulated gene set will then be computationally and experimentally evaluated for regulation of genes in the secondary level of control. In Aim 2, experimental infections with pathogenic bacteria will be used to define the 20E modulation of the immune system in vivo. Both 20E and innate immune signaling are highly conserved across insects, including in agricultural pests and human disease vectors, and the endocrine-mediated development-immunity pleiotropy can be exploited for insect control mechanisms to promote food security and improve public health. More broadly, the principles hormonal regulation of immune function are likely to be conserved across all animals, including humans, and pleiotropic endocrine GRNs allow evolutionary adaptation in life history balances. The methodology and workflow developed in this proposal can be applied broadly to hierarchical gene regulatory networks in any system.