Chromatin swelling drives neutrophil extracellular trap release

Abstract

Neutrophilic granulocytes are able to release their own DNA as neutrophil extracellular traps (NETs) to capture and eliminate pathogens. DNA expulsion (NETosis) has also been documented for other cells and organisms, thus highlighting the evolutionary conservation of this process. Moreover, dysregulated NETosis has been implicated in many diseases, including cancer and inflammatory disorders. During NETosis, neutrophils undergo dynamic and dramatic alterations of their cellular as well as sub-cellular morphology whose biophysical basis is poorly understood. Here we investigate NETosis in real-time on the single-cell level using fluorescence and atomic force microscopy. Our results show that NETosis is highly organized into three distinct phases with a clear point of no return defined by chromatin status. Entropic chromatin swelling is the major physical driving force that causes cell morphology changes and the rupture of both nuclear envelope and plasma membrane. Through its material properties, chromatin thus directly orchestrates this complex biological process.

Nuclear lamin B is crucial to the nuclear envelope integrity and extracellular trap release in neutrophils

“…All of the above results suggest that nuclear lamin B was not destructively cleaved during NETotic nuclear envelope rupture. Therefore, our results are in line with several other studies (Amulic et al, 2017;Neubert et al, 2018) and suggest that the rupture of the nuclear envelope appears to be a distinct process from the previously described lysis or dissolution of the nuclear envelope (Fuchs et al, 2007;Papayannopoulos et al, 2010;Yipp and Kubes, 2013).…”

“…In agreement with our novel findings, a recent work from Zychlinsky and colleagues reported that cyclin dependent kinases (CDK) 4/6 might be important in the regulation of neutrophil NETosis through phosphorylation of lamin A/C (Amulic et al, 2017). In addition, Neubert et al reported that nuclear chromatin swelling provides physical driving force that causes the nuclear envelope rupture and release of neutrophil NETs (Neubert et al, 2018).…”

“…However, all of these latest novel findings from our and other groups (Amulic et al, 2017;Neubert et al, 2018) do not support the lytic hypothesis of nuclear envelope rupture in NETosis, at least in the early stage of this programmed cell death (Neubert et al, 2018).…”

“…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).…”

“…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.…”