Genetic targeting of Card19 is linked to disrupted NINJ1 expression, impaired cell lysis, and increased susceptibility to Yersinia infection

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-deficient mice (Card19lxcn) whose macrophages were protected from cell lysis and showed reduced apoptosis and pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as an independently-generated CRISPR/Cas9 Card19 knockout mouse line (Card19Null) showed no defect in macrophage cell lysis. Notably, Card19 is located on chromosome 13, immediately adjacent to Ninj1, which was recently found to regulate cell lysis downstream of GSDMD activation. RNA-seq and western blotting revealed that Card19lxcn BMDMs have significantly reduced NINJ1 expression, and reconstitution of Ninj1 in Card19lxcn immortalized BMDMs restored their ability to undergo cell lysis in response to caspase-dependent cell death stimuli. Card19lxcn mice exhibited increased susceptibility to Yersinia infection, whereas independently-generated Card19Null mice did not, demonstrating that cell lysis itself plays a key role in protection against bacterial infection, and that the increased infection susceptibility of Card19lxcn mice is attributable to loss of NINJ1. Our findings identify genetic targeting of Card19 being responsible for off-target effects on the adjacent gene Ninj1, disrupting the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage and impacting host survival and bacterial control during Yersinia infection.

Morphological and phylogenetic evidence for two new species of Russula subg. Heterophyllidia from Guangdong Province of China

Two new species of Russula subg. Heterophyllidia from Guangdong Province of China were described and illustrated based on morphological characters, and their identity supported by molecular phylogeny. R. luofuensis is morphologically characterized by a grayish yellow to brownish orange pileus center with a purplish gray to grayish magenta margin, a surface that is cracked and broken into small golden-brown patches, subglobose to broadly ellipsoid basidiospores with warts fused in short or long chains and a suprapellis composed of hyphal extremities with inflated, ellipsoid or globose cells and attenuated terminal cell. R. subbubalina is distinguished by the blanched almond to dark salmon pileus that is cracked with age, subglobose to broadly ellipsoid basidiospores with wart fused in short or long chains and frequently connected by line connections, a suprapellis with hyphal ends composed of inflated or ellipsoid cells and attenuated terminal cell, and pileocystidia that are mainly clavate and sometimes with round or ellipsoid appendage. The phylogenetic analyses based on ITS-nrLSU-mtSSU-TEF1 dataset were performed using maximum likelihood and Bayesian analysis. In terms of morphological features and molecular data, the former species belongs to subsect. Virescentinae, whereas the latter comes under subsect. Heterophyllinae.

Regulators of the secretory pathway have distinct inputs into single-celled branching morphogenesis and seamless tube formation in the Drosophila trachea

Biological tubes serve as conduits through which gas, nutrients and other important fluids are delivered to tissues. Most biological tubes consist of multiple cells connected by epithelial junctions. Unlike these multicellular tubes, seamless tubes are unicellular and lack junctions. Seamless tubes are present in various organ systems, including the vertebrate vasculature, C.elegans excretory system, and Drosophila tracheal system. The Drosophila tracheal system is a network of air-filled tubes that delivers oxygen to all tissues. Specialized cells within the tracheal system, called terminal cells, branch extensively and form seamless tubes. Terminal tracheal tubes are polarized; the lumenal membrane has apical identity whereas the outer membrane exhibits basal characteristics. Although various aspects of membrane trafficking have been implicated in terminal cell morphogenesis, the precise secretory pathway requirements for basal and apical membrane growth have yet to be elucidated. In the present study, we demonstrate that anterograde trafficking, retrograde trafficking and Golgi-to-plasma membrane vesicle fusion are each required for the complex branched architecture of the terminal cell, but their inputs during seamless lumen formation are more varied. The COPII subunit, Sec 31, and ER exit site protein, Sec16, are critical for subcellular tube architecture, whereas the SNARE proteins Syntaxin 5, Syntaxin 1 and Syntaxin15 are required for seamless tube growth and maintenance. These data suggest that distinct components of the secretory pathway have differential contributions to basal and apical membrane growth and maintenance during terminal cell morphogenesis.

Behavioural and ecological data on Dryudella stigma (Panzer, 1809) (Hymenoptera, Astatidae) with the first description of the mature larva

Data on the bionomics of Dryudella stigma (Panzer, 1809) in Poland are presented. The larva is described for the first time for a European Dryudella, and second in the world with only larva of D. immigrans described before. The egg stage lasts 2–3 days, and the larval stage 11–14 days. The praepupal stage lasts the longest time and overwinters in a cocoon from mid-July to the end of May. The female provisions with an average of seven bug nymphs per cell, mainly of Aelia acuminata (Linnaeus, 1758) (Pentatomidae), less often Ceraleptus lividus Stein, 1858 (Coreidae) and Corizus hyoscyami (Linnaeus, 1758) (Rhopalidae). Nests are built in sandy, shaded areas and consist of the main burrow, 10–12 cm long, and one terminal cell. Males are most often found in sunny areas. Adults visit Achillea millefolium L., Asteraceae, as the main source of nectar.

The Card19 locus of murine chromosome 13 regulates terminal cell lysis downstream of caspase activation and Gasdermin-D cleavage

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-/- mice whose macrophages were protected from cell death and showed reduced pore formation during apoptosis or pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as two independently-generated CRISPR/Cas9 Card19 knockout mice showed no defect in macrophage cell lysis. The original Card19-/- line was generated in a 129SvEvBrd background, and SNP analysis revealed a six megabase region of 129 origin co-segregating with the Card19 locus. Card19 is located on chromosome 13, adjacent to Ninj1, which was recently reported to regulate cell lysis downstream of GSDMD activation. Nonetheless, we could not detect major defects in NINJ1 protein expression or mutations in Ninj1 coding sequence in Card19-/- mice. Mice from the original Card19-/- line exhibited significantly increased susceptibility to Yersinia infection, demonstrating that cell lysis itself plays a key role in protection against bacterial infection. Our findings identify a locus on murine chromosome 13 that regulates the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage, and implicates additional NINJ1-independent factors that control terminal cell lysis.

Endothelial cell adhesion and blood response to hemocompatible peptide 1 (HCP-1), REDV, and RGD peptide sequences with free N-terminal amino groups immobilized on a biomedical expanded polytetrafluorethylene surface

Free N-terminal cell adhesive peptides are assessed from the viewpoint of endothelial cell adhesion and blood response.

Coordinated crosstalk between microtubules and actin by a spectraplakin regulates lumen formation and branching

Subcellular lumen formation by single-cells involves complex cytoskeletal remodelling. We have previously shown that centrosomes are key players in the initiation of subcellular lumen formation in Drosophila melanogaster, but not much is known on the what leads to the growth of these subcellular luminal branches or makes them progress through a particular trajectory within the cytoplasm. Here, we have identified that the spectraplakin Short-stop (Shot) promotes the crosstalk between MTs and actin, which leads to the extension and guidance of the subcellular lumen within the tracheal terminal cell (TC) cytoplasm. Shot is enriched in cells undergoing the initial steps of subcellular branching as a direct response to FGF signalling. An excess of Shot induces ectopic acentrosomal luminal branching points in the embryonic and larval tracheal TC leading to cells with extra-subcellular lumina. These data provide the first evidence for a role for spectraplakins in single-cell lumen formation and branching.