Innate immune responses combat infectious microbes by driving inflammation, host-defense pathways and adaptive immunity. Multiple genes within distinct functional categories are coordinately and temporally regulated by transcriptional on and off switches that account for the specificity of gene expression in response to microbial triggers. Well-defined DNA binding transcription factors, transcriptional co-regulators and chromatin modifying complexes collaborate to coordinate transcription of immune genes. Recent evidence indicates that non-coding RNAs also contribute to gene regulation in diverse biological contexts. A growing body of evidence including our own published studies supports their importance in the immune system. lncRNAs have been identified in macrophages, dendritic cells, neutrophils, T-cells and B-cells during their differentiation and/or activation. We have found that these lncRNAs in turn regulate expression of other immune genes. This proposal is based on our hypothesis that lncRNAs are associated with chromatin in immune cells in order to regulate transcription of immune genes. We propose to identify and catalogue chromatin-associated lncRNAs in both resting and activated human macrophages and employ cutting edge molecular approaches to functionally characterize the genomic targets of these regulators. These objectives will be met using two specific aims:
In aim 1 we will identify chromatin-associated lncRNAs (ca-lncRNAs) in an unbiased manner in human macrophages using sucrose density gradient fractionation as well as chromatin Immunoprecipitation coupled to RNA-sequencing. The regulation of these ca-lncRNAs will then be profiled using Nanostring technology to assess the temporal regulation and signaling mechanisms regulating their expression in response to microbial products, viral and bacterial pathogens.
In aim2 we will identify the genomic targets and protein-binding partners of chromatin-associated lncRNAs using RNA antisense purification (RAP) coupled to DNA-Sequencing or Mass Spectrometry, respectively. These studies will define functionally important lncRNAs acting at the level of chromatin to modulate macrophage transcriptional responses and define genomic targets of ca-lncRNAs regulated during host-pathogen interactions. This unbiased identification of functionally relevant lncRNAs will also provide a platform for future studies that characterize these genes in greater detail. They will also provide new strategies to study lncRNA in other biological contexts.

Public Health Relevance

The immune system provides protection against invading pathogens trying to enter the body, but these immune defenses can also be dangerous leading to tissue damage resulting in diseases such as atherosclerosis (heart disease), arthritis and cancer. It is essential that these inflammatory pathways are carefully regulated and kept in check. It is now clear that both proteins as well as RNA molecules can act together to regulate these responses. The major class of RNA molecules produced from the genome is called long noncoding RNA (lncRNA), which appear to regulate the expression of other protein coding genes. The studies proposed in this application are focused on identifying functionally important long-non-coding RNA molecules that are positioned at the genome regulating expression of other genes. By identifying these RNAs and the genes they are bound to we can begin to understand how inflammatory gene expression is regulated. A better understanding of these events is needed to boost immunity for vaccines or to limit immunity to prevent disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AI115448-02
Application #
8966645
Study Section
Cellular and Molecular Immunology - A Study Section (CMIA)
Program Officer
Palker, Thomas J
Project Start
2014-12-01
Project End
2016-11-30
Budget Start
2015-12-01
Budget End
2016-11-30
Support Year
2
Fiscal Year
2016
Total Cost
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
603847393
City
Worcester
State
MA
Country
United States
Zip Code
Elling, Roland; Robinson, Elektra K; Shapleigh, Barbara et al. (2018) Genetic Models Reveal cis and trans Immune-Regulatory Activities for lincRNA-Cox2. Cell Rep 25:1511-1524.e6
Atianand, Maninjay K; Hu, Wenqian; Satpathy, Ansuman T et al. (2016) A Long Noncoding RNA lincRNA-EPS Acts as a Transcriptional Brake to Restrain Inflammation. Cell 165:1672-1685
Elling, Roland; Chan, Jennie; Fitzgerald, Katherine A (2016) Emerging role of long noncoding RNAs as regulators of innate immune cell development and inflammatory gene expression. Eur J Immunol 46:504-12