MUNC13-4 Protein Regulates the Oxidative Response and Is Essential for Phagosomal Maturation and Bacterial Killing in Neutrophils

Monfregola, Jlenia; Johnson, Jack; Meijler, Michaël M.; Napolitano, Gennaro; Catz, Sergio D.

doi:10.1074/jbc.m112.414029

Cited by 40 publications

(89 citation statements)

References 69 publications

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“…However, it has recently been demonstrated that (1) NK cell cytotoxicity has a distinct immunoregulatory role in HLH disease, 19 and (2) Munc13-4 has a critical function in platelets and neutrophils. [20][21][22][23] By using an LV coding for a Munc13-4/GFP fusion protein, our present study demonstrated that gene transfer into murine HSCs resulted in transgene expression in all the hematopoietic lineages, including NK and myeloid cells. Thus, HSC gene correction (providing multilineage Munc13-4 expression) might be a valuable treatment option for FHL3 in patients.…”

Section: Discussionmentioning

confidence: 94%

Gene transfer into hematopoietic stem cells reduces HLH manifestations in a murine model of Munc13-4 deficiency

et al. 2017

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Section: Discussionmentioning

confidence: 94%

Gene transfer into hematopoietic stem cells reduces HLH manifestations in a murine model of Munc13-4 deficiency

et al. 2017

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“…Despite its characterization as a Rab27a effector, Munc13-4 regulates important Rab27a-independent functions including the modulation of the secretion of readily releasable secretory vesicles (12), phagosomal maturation (15), and the differential secretory response to stimuli of lytic granules (16). To determine whether Munc13-4 mediates any regulatory function by binding to Rab GTPases other than Rab27a, we developed a screening approach to identify new binding partners of Munc13-4.…”

Section: Resultsmentioning

confidence: 99%

“…Based on the similarity of this phenotype with that observed in Rab27a-deficient patients along with pull-down studies, protein-protein interaction, and functional assays, Munc13-4 was identified as a Rab27a-specific effector (14). Despite shared roles in regulating common secretory pathways, Munc13-4 regulates many cellular functions in a Rab27a-independent manner including phagosomal maturation in neutrophils (15), the differential secretory response to stimuli in NK cells (16) and the regulation of readily releasable secretory granules in granulocytes (12). It is therefore unclear how these discrete molecular events during exocytosis are regulated by Munc13-4.…”

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confidence: 99%

Munc13-4 Is a Rab11-binding Protein That Regulates Rab11-positive Vesicle Trafficking and Docking at the Plasma Membrane

Johnson¹,

He²,

Ramadass³

et al. 2016

Journal of Biological Chemistry

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The small GTPase Rab11 and its effectors control trafficking of recycling endosomes, receptor replenishment and the up-regulation of adhesion and adaptor molecules at the plasma membrane. Despite recent advances in the understanding of Rab11-regulated mechanisms, the final steps mediating docking and fusion of Rab11-positive vesicles at the plasma membrane are not fully understood. Munc13-4 is a docking factor proposed to regulate fusion through interactions with SNAREs. In hematopoietic cells, including neutrophils, Munc13-4 regulates exocytosis in a Rab27a-dependent manner, but its possible regulation of other GTPases has not been explored in detail. Here, we show that Munc13-4 binds to Rab11 and regulates the trafficking of Rab11-containing vesicles. Using a novel Time-resolved Fluorescence Resonance Energy Transfer (TR-FRET) assay, we demonstrate that Munc13-4 binds to Rab11a but not to dominant negative Rab11a. Immunoprecipitation analysis confirmed the specificity of the interaction between Munc13-4 and Rab11, and super-resolution microscopy studies support the interaction of endogenous Munc13-4 with Rab11 at the single molecule level in neutrophils. Vesicular dynamic analysis shows the common spatio-temporal distribution of Munc13-4 and Rab11, while expression of a calcium binding-deficient mutant of Munc13-4 significantly affected Rab11 trafficking. Munc13-4-deficient neutrophils showed normal endocytosis, but the trafficking, upregulation, and retention of Rab11-positive vesicles at the plasma membrane was significantly impaired. This correlated with deficient NADPH oxidase activation at the plasma membrane in response to Rab11 interference. Our data demonstrate that Munc13-4 is a Rab11-binding partner that regulates the final steps of Rab11-positive vesicle docking at the plasma membrane.Rab GTPases are membrane organizers that regulate the specificity of subcellular compartments and trafficking of these compartments through interactions with specific effectors (1). In particular, Rab11, a ubiquitously expressed GTPase, has been implicated in the regulation of several intracellular mechanisms including recycling endosome trafficking, transport of cargo from the sorting endosomes and the trans-Golgi network to the endosomal recycling compartment and regulation of trafficking during cytokinesis (2). In addition, the Rab11 family of GTPases, comprised by Rab11a, Rab11b and Rab11c/Rab25, are known to regulate several cellular functions including exocytosis (3), phagocytosis (4), and cell-cell communication (5). Rab11 functions are regulated by a broad range of molecular interactors and effectors (2), and proteomic analyses have identified a large array of structurally diverse molecules that interact with Rab11. The most studied family of Rab11 effectors, FIPs (Rab11 family-interacting proteins), 4 comprises five members. Rip11, FIP2, and RCP are characterized by the presence of a conserved Rab11-binding domain at their C terminus and a C2 domain at their N terminus, and FIP3 and FIP4 are characterized by t...

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“…In our study, more than 50% of P. stomatis phagosomes recruited specific granules, as visualized by the granule marker lactoferrin. The lactoferrin-positive integrative pattern observed on P. stomatis phagosomes is an indication of granule fusion (31). Host proteins like lactoferrin can sequester iron, an important bacterial nutritional metal, and prevent microbial growth.…”

Section: Discussionmentioning

confidence: 99%

Peptoanaerobacter stomatis Primes Human Neutrophils and Induces Granule Exocytosis

et al. 2017

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Peptoanaerobacter stomatis is a newly appreciated taxon associated with periodontal diseases; however, little is known about the organism's pathogenic potential or its interaction with the host immune response. Neutrophils are the most abundant innate immune cell present in the gingival tissue and function to constrain the oral microbial challenge. However, some periodontal pathogens have developed strategies to evade phagocytosis and killing by neutrophils. Therefore, to begin to understand the role of P. stomatis in periodontitis, we studied its interactions with human neutrophils. Our data showed that after 30 min of incubation, neutrophils failed to engulf P. stomatis efficiently; however, when P. stomatis was internalized, it was promptly eradicated. P. stomatis challenge induced a robust intracellular respiratory burst; however, this response did not contribute to bacterial killing. Minimal superoxide release was observed by direct bacterial challenge; however, P. stomatis significantly increased N-formyl-methionyl-leucyl phenylalanine (fMLF)-stimulated superoxide release to an extent similar to that of cells primed with tumor necrosis factor alpha (TNF-␣). When neutrophils were challenged with P. stomatis, 52% of the bacterium-containing phagosomes were enriched for the specific granule marker lactoferrin and 82% with the azurophil granule marker elastase. P. stomatis challenge stimulated exocytosis of the four neutrophil granule subtypes. Moreover, P. stomatis susceptibility to extracellular killing could be attributed to the exocytosis of antimicrobial components present in neutrophil granules. Priming neutrophils for an enhanced respiratory burst together with promoting granule content release could contribute to the chronic inflammation and tissue destruction that characterize periodontal diseases.

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MUNC13-4 Protein Regulates the Oxidative Response and Is Essential for Phagosomal Maturation and Bacterial Killing in Neutrophils

Cited by 40 publications

References 69 publications

Gene transfer into hematopoietic stem cells reduces HLH manifestations in a murine model of Munc13-4 deficiency

Gene transfer into hematopoietic stem cells reduces HLH manifestations in a murine model of Munc13-4 deficiency

Munc13-4 Is a Rab11-binding Protein That Regulates Rab11-positive Vesicle Trafficking and Docking at the Plasma Membrane

Peptoanaerobacter stomatis Primes Human Neutrophils and Induces Granule Exocytosis

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