In response to a variety of stimuli, human neutrophils undergo a respiratory burst in which oxygen is converted to superoxide (O2-), hydrogen peroxide, hypochlorous acid, and possibly hydroxyl radicals. While these agents generally serve a beneficial role in killing pathogenic microbes, they also can cause serious damage to normal tissues during inflammation. In addition, several human diseases are associated with a failure to regulate appropriately the production of these reactive oxygen derivatives. In the adult respiratory distress syndrome, for example, pulmonary damage is thought to occur because of excessive production of phagocyte oxidants, while in chronic granulomatous disease (CGD), failure to produce these molecules leads to life-threatening infections. The studies proposed are part of a long-range goal to elucidate the biochemical mechanisms that regulate the enzyme responsible for generating reactive oxygen derivatives in phagocytes, NADPH oxidase. A second long-range goal is to develop drugs capable of modulating the level of activity of this enzyme. In clinical situations characterized by excessive inflammation, pharmacologic agents Out down-regulate oxidase activity might provide therapeutic benefit. Conversely, drugs capable of enhancing its activity in such a way that the antimicrobial efficiency of the phagocyte is improved might be of benefit to patients suffering from overwhelming infections or from various types of immunodeficiencies. Several key experimental methods have been developed in this laboratory for studying this problem. A fully soluble cell-free oxidase activation system has been developed, characterized, and optimized. A group of 60 CGD patients who have a variety of different mutations encompassing four of the oxidase components have now been characterized and are regularly available for further studies. Through the use of the fully soluble cell-free system, it has been possible to reconstitute oxidase activity in both the defective membranes and cytosols from these patients using partially purified oxidase components. The following specific aims are designed to build upon this progress and focus on several of the major technical and conceptual deficiencies that currently impede the achievement of the long-range goals of this project: 1) to identify and purify the remaining cytosolic oxidase components and study their intermolecular interactions in unstimulated, primed, and activated normal and CGD neutrophils; 2) to identify and purify remaining membrane oxidase components and study their molecular interactions both within the membrane and with cytosolic components; 3) to determine the role of GTP, low molecular weight GTP-binding proteins, ATP, and lipid second messengers in the regulation of NADPH oxidase activation in the cell-free system. These studies may ultimately lead to new pharmacologic methods for controlling oxygen radical-mediated tissue damage as well as to an improved understanding of the pathophysiology of immune disorders such as CGD.
| von Löhneysen, Katharina; Noack, Deborah; Wood, Malcolm R et al. (2010) Structural insights into Nox4 and Nox2: motifs involved in function and cellular localization. Mol Cell Biol 30:961-75 |
| Luxen, Sylvia; Noack, Deborah; Frausto, Monika et al. (2009) Heterodimerization controls localization of Duox-DuoxA NADPH oxidases in airway cells. J Cell Sci 122:1238-47 |
| Lehmann, Mandy; Noack, Deborah; Wood, Malcolm et al. (2009) Lung epithelial injury by B. anthracis lethal toxin is caused by MKK-dependent loss of cytoskeletal integrity. PLoS One 4:e4755 |
| von Lohneysen, Katharina; Noack, Deborah; Jesaitis, Algirdas J et al. (2008) Mutational analysis reveals distinct features of the Nox4-p22 phox complex. J Biol Chem 283:35273-82 |
| Pacquelet, Sandrine; Lehmann, Mandy; Luxen, Sylvia et al. (2008) Inhibitory action of NoxA1 on dual oxidase activity in airway cells. J Biol Chem 283:24649-58 |
| Maroto, B; Ye, M B; von Lohneysen, K et al. (2008) P21-activated kinase is required for mitotic progression and regulates Plk1. Oncogene 27:4900-8 |
| Luxen, Sylvia; Belinsky, Steven A; Knaus, Ulla G (2008) Silencing of DUOX NADPH oxidases by promoter hypermethylation in lung cancer. Cancer Res 68:1037-45 |
| Zhu, Yanmin; Marchal, Christophe C; Casbon, Amy-Jo et al. (2006) Deletion mutagenesis of p22phox subunit of flavocytochrome b558: identification of regions critical for gp91phox maturation and NADPH oxidase activity. J Biol Chem 281:30336-46 |
| Zhu, Hao; Larade, Kevin; Jackson, Timothy A et al. (2004) NCB5OR is a novel soluble NAD(P)H reductase localized in the endoplasmic reticulum. J Biol Chem 279:30316-25 |
| Heyworth, Paul G; Cross, Andrew R; Curnutte, John T (2003) Chronic granulomatous disease. Curr Opin Immunol 15:578-84 |
Showing the most recent 10 out of 100 publications
