Neutrophils contain four types of secretory organelles that hold specialized proteins essential for their microbicidal activity. Unrestricted release of the toxic content of their granules would be injurious to the host. Exocytosis of these granules is hierarchical and differentially regulated. This correlates with the role the cargo proteins play in the processes of adhesion, migration, chemotaxis, phagocytosis and production of reactive oxygen species. The mechanism of vesicle exocytosis in neutrophils is poorly understood. Rab27a is a small GTPase associated with secretory processes. Its role in granulocytes is unknown. Genetic defects in Rab27a are associated with the human immunodeficiency Griscelli syndrome in which neutrophil function may be altered. On the basis of extensive evidence from our laboratory, we propose the following hypotheses: 1) Rab27a and its effectors JFC1 and Munc13-4 play a central role in exocytosis in neutrophils and in associated functions that depend on granular protein mobilization;2) granules implicated in cargo release into the surrounding milieu are molecularly and mechanistically different from those involved in cargo release into the phagosome;3) JFC1 provides the specificity required to target Rab27a-containing vesicles to the docking site at the plasma membrane during exocytosis while Munc13-4 regulates the fusion process. To understand the mechanisms of Rab27a-dependent vesicular trafficking in granulocytes, we propose three specific aims: 1) To examine the role that Rab27a plays in exocytosis and phagocytosis in granulocytes, using Rab27a-deficient mice;2) To identify the mechanisms that regulate the docking of Rab27a-containing vesicles in granulocytes by electron and evanescence fluorescence microscopy using JFC1-deficient cells;3) To identify the cellular and biochemical events regulated by the Rab27a-effector Munc13-4. We plan to elucidate the mechanism underlying vesicle fusion and exocytosis using neutrophils from Munc13-4-deficient mice. The results of the proposed research plan should uncover the molecular mechanisms of vesicle docking, fusion and secretion in neutrophils and lead to effective molecular strategies for the treatment of inflammatory diseases.
Neutrophils are white cells that circulate in the blood. They play an essential role in protecting us from microbial infections through the production and release of substances that are toxic to the microorganisms. These toxic products, when released in large amounts or in an uncontrolled manner, can be harmful to the host (humans). The study of the mechanism that controls the release of toxic products by neutrophils will lead to the design of therapeutic strategies for the control of diseases like arthritis and other inflammatory disorders.
|Catz, Sergio D (2014) The role of Rab27a in the regulation of neutrophil function. Cell Microbiol 16:1301-10|
|Catz, Sergio Daniel (2013) Regulation of vesicular trafficking and leukocyte function by Rab27 GTPases and their effectors. J Leukoc Biol 94:613-22|
|Johnson, Jennifer L; Napolitano, Gennaro; Monfregola, Jlenia et al. (2013) Upregulation of the Rab27a-dependent trafficking and secretory mechanisms improves lysosomal transport, alleviates endoplasmic reticulum stress, and reduces lysosome overload in cystinosis. Mol Cell Biol 33:2950-62|
|Johnson, Jennifer L; Monfregola, Jlenia; Napolitano, Gennaro et al. (2012) Vesicular trafficking through cortical actin during exocytosis is regulated by the Rab27a effector JFC1/Slp1 and the RhoA-GTPase-activating protein Gem-interacting protein. Mol Biol Cell 23:1902-16|
|Johnson, Jennifer L; Hong, Hong; Monfregola, Jlenia et al. (2011) Increased survival and reduced neutrophil infiltration of the liver in Rab27a- but not Munc13-4-deficient mice in lipopolysaccharide-induced systemic inflammation. Infect Immun 79:3607-18|
|Johnson, Jennifer L; Hong, Hong; Monfregola, Jlenia et al. (2011) Munc13-4 restricts motility of Rab27a-expressing vesicles to facilitate lipopolysaccharide-induced priming of exocytosis in neutrophils. J Biol Chem 286:5647-56|
|Johnson, Jennifer L; Brzezinska, Agnieszka A; Tolmachova, Tanya et al. (2010) Rab27a and Rab27b regulate neutrophil azurophilic granule exocytosis and NADPH oxidase activity by independent mechanisms. Traffic 11:533-47|
|Munafo, Daniela B; Johnson, Jennifer L; Brzezinska, Agnieszka A et al. (2009) DNase I inhibits a late phase of reactive oxygen species production in neutrophils. J Innate Immun 1:527-42|