No abstract

It's not clear how nuclear envelope (NE) is ruptured for chromatin externalization duringNETosis. The membrane rupture during neutrophil NET release was described as a membrane lysis process, this notion, however, has been questioned. Here, we found that lamin B, the structural NE component, was involved in NETosis. Unexpectedly, lamin B was not fragmented by destructive proteolysis, but rather disassembled into its intact full-length molecule, in NETotic cells with ruptured NE. In the mechanistic study, our experiments demonstrated that cytosolic PKCα translocated to the nucleus, where it serves as a NETotic lamin kinase to induce lamin B phosphorylation, following by lamina disassembly and NE rupture. To determine causality, we found that decreasing lamin B phosphorylation, by PKCα inhibition or genetic deletion, or mutation at the PKCα consensus phosphorylation sites of lamin B, attenuated extracellular trap formation. Importantly, strengthening NE by lamin B overexpression attenuated neutrophil NETosis in vivo and alleviated exhibition of NET-associated inflammatory cytokines in UVB irradiated skin of lamin B transgenic mice. These findings advance our understanding of NETosis process and elucidate a cellular mechanism that PKCα-mediated lamin B phosphorylation drives nuclear envelope rupture for NET release in neutrophils. Papayannopoulos et al., 2010), while neutrophil elastase is dispensable for NET formation as neutrophils from neutrophil elastase deficient mice could still undergo NETosis in a recent study (Martinod et al., 2016).Release of the decondensed nuclear chromatin, through the broken nuclear and plasma membranes, is the key step for formation of the neutrophil NET backbone. Neutrophil NET release was described as a lytic cell death mechanism (Fuchs et al., 2007;Papayannopoulos et al., 2010;Yipp and Kubes, 2013). However, this notion has been questioned by several recent observations (Pilsczek et al., 2010;Amulic et al., 2017;Neubert et al., 2018). Rupture of the nuclear envelope/nuclear membrane appears to be a distinct process from the previously described lysis or dissolution of the nuclear envelope (Amulic et al., 2017;Neubert et al., 2018).Since nuclear chromatin is enclosed by the nuclear envelope, its breakdown is, therefore, a prerequisite, and a key step, for externalization of nuclear chromatin and extracellular trap formation. (Papayannopoulos et al., 2010;Yipp and Kubes, 2013) However, the relevant mechanisms that regulate nuclear envelope rupture in NETosis are still unclear.The nuclear envelope consists of outer and inner lipid nuclear membranes (ONM and INM) and nuclear lamina, a protein filament meshwork that provides structural scaffold to reinforce nuclear envelope integrity (Goldberg et al., 2008;Schreiber and Kennedy, 2013;Stewart et al., 2007).Nuclear lamins are categorized as either A-type (A, C) or B-type (B1, B2) lamins (Schreiber and Kennedy, 2013). A-type lamins form thick filament bundles that provide mechanical rigidity to the nucleus (Goldberg et al., 2008;Lammerding...