Identifying novel transcription factors involved in the inflammatory response by using binding site motif scanning in genomic regions defined by histone acetylation.

Peter S Askovich,Stephen A Ramsey,Alan Aderem,Theo A Knijnenburg,Kathleen A Kennedy,Alan H Diercks

doi:10.1371/journal.pone.0184850

Abstract

The innate immune response to pathogenic challenge is a complex, multi-staged process involving thousands of genes. While numerous transcription factors that act as master regulators of this response have been identified, the temporal complexity of gene expression changes in response to pathogen-associated molecular pattern receptor stimulation strongly suggest that additional layers of regulation remain to be uncovered. The evolved pathogen response program in mammalian innate immune cells is understood to reflect a compromise between the probability of clearing the infection and the extent of tissue damage and inflammatory sequelae it causes. Because of that, a key challenge to delineating the regulators that control the temporal inflammatory response is that an innate immune regulator that may confer a selective advantage in the wild may be dispensable in the lab setting. In order to better understand the complete transcriptional response of primary macrophages to the bacterial endotoxin lipopolysaccharide (LPS), we designed a method that integrates temporally resolved gene expression and chromatin-accessibility measurements from mouse macrophages. By correlating changes in transcription factor binding site motif enrichment scores, calculated within regions of accessible chromatin, with the average temporal expression profile of a gene cluster, we screened for transcriptional factors that regulate the cluster. We have validated our predictions of LPS-stimulated transcriptional regulators using ChIP-seq data for three transcription factors with experimentally confirmed functions in innate immunity. In addition, we predict a role in the macrophage LPS response for several novel transcription factors that have not previously been implicated in immune responses. This method is applicable to any experimental situation where temporal gene expression and chromatin-accessibility data are available.

Highlights

Macrophages are long-lived coordinating cells of the innate immune system
One of the key chromatin marks directing the transcriptional response to endotoxin stimulation in macrophages is histone acetylation (HAc), which is associated with open chromatin and active promoters [10, 12]
Functional transcription factors (TFs) binding sites (TFBS) are often found within regions of histone acetylation, and our previous work has shown that the binding sites within histone-acetylated regions tend to appear as distinct features in the quantitative signal that represents the local amount of HAc ChIP-seq fragment recovery [13]

Summary

Introduction

Activation of tissue macrophages by Toll-like receptor (TLR) stimulation initiates a dynamic program of gene expression changes involving hundreds of genes that are associated with processes such as phagocytosis, antigen presentation, immunoregulation, and non-oxidative metabolism [1,2,3,4] This gene expression program involves scores of transcription factors (TFs) whose activation is regulated both hierarchically [5,6,7] and temporally [7,8,9] and whose accessible binding sites in the genome change over time due to stimulation-dependent alterations in epigenetic state of the chromatin [7, 10, 11]. Mammalian TF binding sites are often distal to the transcription start site (TSS), for example, in enhancers that can be many kilobase distant [11, 13, 27, 28]

Methods

Results

Conclusion