This is a competitive renewal entitled, """"""""Enzymes modulating second messengers in neutrophils"""""""", requesting five more years of finding, for years 10-14. (adapted from the applicant's abstract) The objective of this proposal is to define the exact sequence of molecular events that are involved in the stimulation of phagocytic leukocytes. Knowledge gained from these studies will likely to lead to novel strategies for treating infectious diseases and inflammation. This project will now focus on two distinct but critical reactions in the stimulation of neutrophils: (1) the dephosphorylation/activation of cofilin, and (2) the role of different isozyme of protein kinase C (PKC) in the activation of the NADPH oxidase complex which triggers superoxide generation. The investigator and collaborators have recently reported that cofilin undergoes rapid dephosphorylation/activation in stimulated neutrophils Cofilin is an essential actin depolymerizing agent that is critically involved in regulating the actin cytoskeleton, particularly the functional responses of neutrophils, e.g., chemotaxis, phagosome formation and degranulation. Moreover, the investigator has presented evidence that cofilin is regulated by a novel cycle of phosphorylation and dephosphorylation in neutrophils. The nature of the kinases and phosphatase(s) that participate in this cycle are unknown. Specifically, this project focuses on four unexplored areas (five aims) in the signal transduction pathways of neutrophils. These are (1) identifying and characterizing the protein kinase (aim 1) and phosphatase (aims 2) that catalyze the phosphorylation and dephosphorylation of cofiling in neutrophils, (2) elucidating the regulatory mechanisms that trigger dephosphorylation/activation of cofiling during cell stimulation (aim 3), (3) continue investigating the role of PKC in superoxide production with each of the purified isozymes of PKC that are present in neutrophils and their recombinant substrates, i.e., the oxidase subunits p47-phox and p67- phox (aim 4), and (4) monitoring the assembly of the superoxide generating system by immunofluorescence techniques and characterizing interactions between this complex and the cytoskeleton (aim 5). Co-localization of specific isozymes of PKC with the oxidase subunits will be bought. The goal is to forge a solid line between the regulatory properties of the isolated enzymes and control of the stimulus-response phenomena in phagocytic leukocytes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI023323-10
Application #
2688487
Study Section
Special Emphasis Panel (ZRG5-BM-2 (03))
Project Start
1986-08-01
Project End
1999-07-31
Budget Start
1998-08-01
Budget End
1999-07-31
Support Year
10
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Boston Biomedical Research Institute
Department
Type
DUNS #
058893371
City
Watertown
State
MA
Country
United States
Zip Code
02472
Ohira, Taisuke; Bannenberg, Gerard; Arita, Makoto et al. (2004) A stable aspirin-triggered lipoxin A4 analog blocks phosphorylation of leukocyte-specific protein 1 in human neutrophils. J Immunol 173:2091-8
Robinson, John M; Ohira, Taisuke; Badwey, John A (2004) Regulation of the NADPH-oxidase complex of phagocytic leukocytes. Recent insights from structural biology, molecular genetics, and microscopy. Histochem Cell Biol 122:293-304
Ohira, Taisuke; Zhan, Qian; Ge, Qingyuan et al. (2003) Protein phosphorylation in neutrophils monitored with phosphospecific antibodies. J Immunol Methods 281:79-94
Zhan, Qian; Ge, Qingyuan; Ohira, Taisuke et al. (2003) p21-activated kinase 2 in neutrophils can be regulated by phosphorylation at multiple sites and by a variety of protein phosphatases. J Immunol 171:3785-93
Zhan, Qian; Bamburg, James R; Badwey, John A (2003) Products of phosphoinositide specific phospholipase C can trigger dephosphorylation of cofilin in chemoattractant stimulated neutrophils. Cell Motil Cytoskeleton 54:1-15
Oyaizu, Kosuke; Kantarci, Alpdogan; Maeda, Hiroshi et al. (2003) Identification of mRNAs for the various diacylglycerol kinase isoforms in neutrophils from patients with localized aggressive periodontitis. J Periodontal Res 38:488-95
Robinson, John M; Badwey, John A (2002) Rapid association of cytoskeletal remodeling proteins with the developing phagosomes of human neutrophils. Histochem Cell Biol 118:117-25
Lian, Jian P; Crossley, Lisa; Zhan, Qian et al. (2002) The P21-activated protein kinases (Paks) receive and integrate messages from a variety of signaling pathways. Adv Exp Med Biol 507:497-502
Lian, J P; Crossley, L; Zhan, Q et al. (2001) Antagonists of calcium fluxes and calmodulin block activation of the p21-activated protein kinases in neutrophils. J Immunol 166:2643-50
De Mesquita, D D; Zhan, Q; Crossley, L et al. (2001) p90-RSK and Akt may promote rapid phosphorylation/inactivation of glycogen synthase kinase 3 in chemoattractant-stimulated neutrophils. FEBS Lett 502:84-8

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