Computational Identification of Transcriptional Regulators in Human Endotoxemia Tung T. Nguyen 1 , Panagiota T. Foteinou 2 , Steven E. Calvano 3 , Stephen F. Lowry 3 , Ioannis P. Androulakis 2,3 * 1 BioMaPS Institute for Quantitative Biology, Rutgers University, Piscataway, New Jersey, United States of America, 2 Department of Biomedical Engineering, Rutgers University, Piscataway, New Jersey, United States of America, 3 Department of Surgery, Robert Wood Johnson Medical School, University of Medicine and Dentistry, New Jersey, New Brunswick, New Jersey, United States of America Abstract One of the great challenges in the post-genomic era is to decipher the underlying principles governing the dynamics of biological responses. As modulating gene expression levels is among the key regulatory responses of an organism to changes in its environment, identifying biologically relevant transcriptional regulators and their putative regulatory interactions with target genes is an essential step towards studying the complex dynamics of transcriptional regulation. We present an analysis that integrates various computational and biological aspects to explore the transcriptional regulation of systemic inflammatory responses through a human endotoxemia model. Given a high-dimensional transcriptional profiling dataset from human blood leukocytes, an elementary set of temporal dynamic responses which capture the essence of a pro-inflammatory phase, a counter-regulatory response and a dysregulation in leukocyte bioenergetics has been extracted. Upon identification of these expression patterns, fourteen inflammation-specific gene batteries that represent groups of hypothetically ‘coregulated’ genes are proposed. Subsequently, statistically significant cis-regulatory modules (CRMs) are identified and decomposed into a list of critical transcription factors (34) that are validated largely on primary literature. Finally, our analysis further allows for the construction of a dynamic representation of the temporal transcriptional regulatory program across the host, deciphering possible combinatorial interactions among factors under which they might be active. Although much remains to be explored, this study has computationally identified key transcription factors and proposed a putative time-dependent transcriptional regulatory program associated with critical transcriptional inflammatory responses. These results provide a solid foundation for future investigations to elucidate the underlying transcriptional regulatory mechanisms under the host inflammatory response. Also, the assumption that coexpressed genes that are functionally relevant are more likely to share some common transcriptional regulatory mechanism seems to be promising, making the proposed framework become essential in unravelling context-specific transcriptional regulatory interactions underlying diverse mammalian biological processes. Citation: Nguyen TT, Foteinou PT, Calvano SE, Lowry SF, Androulakis IP (2011) Computational Identification of Transcriptional Regulators in Human Endotoxemia. PLoS ONE 6(5): e18889. doi:10.1371/journal.pone.0018889 Editor: Christian Scho ¨ nbach, Kyushu Institute of Technology, Japan Received November 26, 2010; Accepted March 23, 2011; Published May 27, 2011 Copyright: ß 2011 Nguyen et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: TTN, PTF and IPA acknowledge the support from the National Institutes of Health under GM082974. SEC and SFL acknowledge support from the the National Institutes of Health under grant GM34695. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: yannis@rci.rutgers.edu Introduction Inflammation and activation of innate immunity are essential defense responses against invading pathogens and endogenous danger signals. The innate immune response involves the initial recognition of conserved pathogen-associated molecular patterns by members of the Toll-like receptor (TLR) family [1]. The exposure of the host to gram negative bacteria, simulated by lipopolysaccharide (LPS) recognized by TLR-4, triggers intracel- lular signalling cascades which eventually release a lot of pro- and anti- inflammatory cytokines [2]. While the host inflammatory response is essential to resolve the infection or repair the damage and restore the system homeostasis, it also plays a central pathogenic role in a wide spectrum of diseases including sepsis [3]. Under healthy circumstances, inflammatory responses are activated, clear the pathogen in the case of infection, initialize a repair process and then abate [4]. However when anti-inflamma- tory processes fail, an amplified inflammation can turn what is normally a beneficial reparative process into a detrimental physiological state with severe, uncontrolled systemic inflamma- tion [5]. Studies involving experimental human endotoxemia have reported rapid intravenous infusion in doses of 2–4 ng/kg body weight, which effectively induces an acute systemic inflammatory condition that mimics the early flow phase of injury and infection [6,7,8,9,10]. In human peripheral blood leukocytes, intravenous administration of endotoxin elicits dynamic and reproducible changes in the circulating leukocyte population as well as significant changes in blood leukocyte gene expression patterns [11]. This perturbation of leukocyte gene expression involves several thousands of transcripts and accompanies the systemic physiological responses during inflammation, which peaks ,4– 6 hours after endotoxin exposure and resolves within 24 hours, compatible with a large and dynamic regulatory network. PLoS ONE | www.plosone.org 1 May 2011 | Volume 6 | Issue 5 | e18889