Scientific Report 2005

Immunology

Regulation of the Innate Immune Response in Inflammation and Infection

U.G. Knaus, A. Bamberg, M. Lehmann, K. von Loehneysen, S. Luxen, S. Pacquelet, M. Ruse, M. Valo, M. Ye

Innate immune cells are the first line of defense in the fight against invading pathogens. We focus primarily on understanding molecular mechanisms that phagocytes and the pulmonary epithelium use to protect the host from the injury and how some responses wind up damaging the host. For example, second messengers such as reactive oxygen species (ROS) or nitric oxide that are produced during infection can have beneficial as well as detrimental effects. The overall outcome depends on precise spatial and temporal regulation of these second messengers by the affected cell populations. The intracellular signaling pathways that control these turn on–turn off mechanisms are an ideal target for intervention in disease.

Almost all of the processes connected to pathogen uptake, pathogen elimination, and sustained inflammation are governed by small GTPases of the Ras superfamily. Our research centers on the Rho GTPases Rac, Cdc42, and Rho, which are essential regulators for various leukocyte functions ranging from production of ROS to chemotaxis and phagocytosis. Generation of superoxide anion is accomplished by a Rac-dependent NADPH oxidase (Nox) upon stimulation with chemotactic factors or phagocytic stimuli. We have identified several Rac effector protein kinases, p21-activated kinases (PAKs), in leukocytes, and we are investigating the role of PAKs in the generation of superoxide anion. Additionally, we identified and are characterizing downstream targets of PAKs, which are relevant for innate immune cell functions.

GTPases of the Rho family are also involved in signaling cascades, which originate from pathogen-activated Toll-like receptors. Toll-like receptors 2 and 4, stimulated by microbial products derived from gram-positive and gram-negative bacteria, activate Rac1 and RhoA, which initiate 2 independent pathways required for RelA transactivation and subsequent NF-κB–dependent gene transcription. We are studying different aspects of signaling by Toll-like receptors in several primary human cell types, including monocytes and neutrophils, and genetically altered mouse models and the impact of this signaling on innate immune cell functions such as apoptosis and upregulation of proinflammatory mediators.

Another area of research is the interaction and communication between innate immune cells and the pulmonary epithelium. To this end, we established an in vitro reconstitution system for lung epithelium that we use to examine signaling mechanisms initiated by pathogens (Fig. 1).

Fig. 1. Transmission electron micrograph of a 3-dimensional culture of human airway epithelium grown in air-liquid interface culture for 33 days (3900X).

The differentiated and fully functional lung epithelium also serves as a model for studies of lung barrier function and the influence of bacteria-derived ligands and toxins on transmigration of neutrophils. In addition, we will investigate processes leading to uptake of pathogens or environmental particles and the impact of these pathogens on airway epithelial functions.Recently, ROS-generating Nox proteins have been identified in epithelial cells, and work is in progress to study the molecular basis for ROS generation by these novel proteins. Nox proteins may serve as compartmentalized signaling modules, thereby activating or inhibiting signaling cascades via superoxide, or as an epithelial host defense mechanism via hydrogen peroxide–generating Nox/Duox isoforms. Because of their tissue-specific distribution and distinct localization patterns, Nox proteins might have highly specialized functions and undergo isoform-dependent regulation. For example, Nox4, an oxidase expressed in colon tissue and melanomas, is constitutively active in certain conditions and does not require any of the known oxidase components for superoxide generation. Elucidating physiologic stimuli and control mechanisms for these Nox proteins combined with structure-function studies will help define the biological functions of Nox in health and disease.

Publications

Chan, A.Y., Coniglio, S.J., Chuang, Y.Y., Michaelson, D., Knaus, U.G., Philips, M.R., Symons, M. Roles of the Rac1 and Rac3 GTPases in human tumor cell invasion. Oncogene, in press.

Martyn, K.D., Frederick, L.M., von Loehneysen, K., Dinauer, M.C., Knaus, U.G. Functional analysis of Nox4 reveals unique characteristics compared to other NADPH oxidases. Cell. Signal., in press.

Martyn, K.D., Kim, M.J., Quinn, M.T., Dinauer, M.C., Knaus, U.G. p-21 Activated kinase (Pak) regulates NADPH oxidase activation in human neutrophils. Blood, in press.

Yamauchi, A., Marchal, C.C., Molitoris, J., Pech, N., Knaus, U., Towe, J., Atkinson, S.J., Dinauer, M.C. Rac GTPase isoform-specific regulation of NADPH oxidase and chemotaxis in murine neutrophils in vivo: role of the C-terminal polybasic domain. J. Biol. Chem. 280:953, 2005.