A Myeloperoxidase-Containing Complex Regulates Neutrophil Elastase Release and Actin Dynamics during NETosis

Abstract

SummaryNeutrophils contain granules loaded with antimicrobial proteins and are regarded as impermeable organelles that deliver cargo via membrane fusion. However, during the formation of neutrophil extracellular traps (NETs), neutrophil elastase (NE) translocates from the granules to the nucleus via an unknown mechanism that does not involve membrane fusion and requires reactive oxygen species (ROS). Here, we show that the ROS triggers the dissociation of NE from a membrane-associated complex into the cytosol and activates its proteolytic activity in a myeloperoxidase (MPO)-dependent manner. In the cytosol, NE first binds and degrades F-actin to arrest actin dynamics and subsequently translocates to the nucleus. The complex is an example of an oxidative signaling scaffold that enables ROS and antimicrobial proteins to regulate neutrophil responses. Furthermore, granules contain protein machinery that transports and delivers cargo across membranes independently of membrane fusion.

Biology, role and therapeutic potential of circulating histones in acute inflammatory disorders

“…Neutrophil azoruphilic granules contain neutrophil elastase (NE), proteinase 3 (PR3) and cathepsin G (CG); however, only NE is translocated to the nucleus and neither inhibition of PR3 nor CG can prevent this translocation 7. Furthermore, the process does not appear to be mediated by fusion of granules with the nucleus, but rather NE dissociates from the granular membrane in a ROS‐dependent manner, before degrading cytosolic actin, arresting actin dynamics and translocating across the nuclear membrane using specific translocation mechanisms 67. Binding of nucleic acid by proteases initiates a process of degradation of nuclear binding proteins68 and controlled integration of MPO into the forming NET.…”

Characterization of Filifactor alocis and its immune evasion strategies employed against human neutrophils.

“…First, neutrophil elastase (NE), a neutrophil-specific serine protease, functions in histone degradation, promotion of chromatin decondensation in the neutrophil nucleus, degradation of the nuclear envelope, antimicrobial activities within the neutrophil phagosome (170,376,377). Studies performed with both NE and MPO knockout mice show failed induction of NETs and an increased susceptibility to infection (170).…”

“…Studies performed with both NE and MPO knockout mice show failed induction of NETs and an increased susceptibility to infection (170). Tightly associated with NE is myeloperoxidase, which is necessary for NE, as it functions in conjunction with the ROS to translocate NE to the neutrophil nucleus (376). MPO is an enzyme that works to consume hydrogen peroxide in order to form hypochlorous acid (HOCl) among other important anti-microbial oxidants (376).…”

“…Tightly associated with NE is myeloperoxidase, which is necessary for NE, as it functions in conjunction with the ROS to translocate NE to the neutrophil nucleus (376). MPO is an enzyme that works to consume hydrogen peroxide in order to form hypochlorous acid (HOCl) among other important anti-microbial oxidants (376). There are four core histones in NETs (H2A, H2B, H3 and H4), which are responsible for the majority of their protein mass and serve as potent antimicrobials (168).…”

“…Several studies have highlighted the importance of NET component neutrophil elastase (NE) to the effective formation and function of NETs (170,376,377). Therefore, using a kit to detect extracellular release of NE, we sought to determine if its presence was found with nonopsonized and opsonized F. alocis challenge of neutrophils.…”