Search

Given the aggressive spread of COVID-19-related deaths, there is an urgent public health need to support the development of vaccine candidates to rapidly improve the available control measures against SARS-CoV-2. To meet this need, we are leveraging our existing vaccine platform to target SARS-CoV-2. Here, we generated cellular heat shock chaperone protein, glycoprotein 96 (gp96), to deliver SARS-CoV-2 protein S (spike) to the immune system and to induce cell-mediated immune responses. We showed that our vaccine platform effectively stimulates a robust cellular immune response against protein S. Moreover, we confirmed that gp96-Ig, secreted from allogeneic cells expressing full-length protein S, generates powerful, protein S polyepitope-specific CD4+ and CD8+ T cell responses in both lung interstitium and airways. These findings were further strengthened by the observation that protein-S -specific CD8+ T cells were induced in human leukocyte antigen HLA-A2.1 transgenic mice thus providing encouraging translational data that the vaccine is likely to work in humans, in the context of SARS-CoV-2 antigen presentation. [ABSTRACT FROM AUTHOR]

Perforin-2 (P-2) is an antimicrobial protein with unique properties to kill intracellular bacteria. Gamma delta (GD) T cells, as the major T cell population in epithelial tissues, play a central role in protective and pathogenic immune responses in the skin. However, the tissue-specific mechanisms that control the innate immune response and the effector functions of GD T cells, especially the cross-talk with commensal organisms, are not very well understood. We hypothesized that the most prevalent skin commensal microorganism, Staphylococcus epidermidis , may play a role in regulating GD T cell-mediated cutaneous responses. We analyzed antimicrobial protein P-2 expression in human skin at a single cell resolution using an amplified fluorescence in situ hybridization approach to detect P-2 mRNA in combination with immunophenotyping. We show that S. epidermidis activates GD T cells and upregulates P-2 in human skin ex vivo in a cell-specific manner. Furthermore, P-2 upregulation following S. epidermidis stimulation correlates with increased ability of skin cells to kill intracellular Staphylococcus aureus. Our findings are the first to reveal that skin commensal bacteria induce P-2 expression, which may be utilized beneficially to modulate host innate immune responses and protect from skin infections. [ABSTRACT FROM AUTHOR]

Gamma delta (GD) T cells are an unconventional T cell type present in both the epidermis and the dermis of human skin. They are critical to regulating skin inflammation, wound healing, and anti-microbial defense. Similar to CD8+ cytotoxic T cells expressing an alpha beta (AB) TCR, GD T cells have cytolytic capabilities. They play an important role in elimination of cutaneous tumors and virally infected cells and have also been implicated in pathogenicity of several autoimmune diseases. T cell cytotoxicity is associated with the expression of the pore forming protein Perforin. Perforin is an innate immune protein containing a membrane attack complex perforin-like (MACPF) domain and functions by forming pores in the membranes of target cells, which allow granzymes and reactive oxygen species to enter the cells and destroy them. Perforin-2, encoded by the gene MPEG1 , is a newly discovered member of this protein family that is critical for clearance of intracellular bacteria. Cutaneous GD T cells express both Perforin and Perforin-2, but many questions remain regarding the role that these proteins play in GD T cell mediated cytotoxicity against tumors and bacterial pathogens. Here, we review what is known about Perforin expression by skin GD T cells and the mechanisms that contribute to Perforin activation. [ABSTRACT FROM AUTHOR]

Perforin‐2 (P‐2) is a recently described antimicrobial protein with unique properties to kill intracellular bacteria. We investigated P‐2 expression pattern and cellular distribution in human skin and its importance in restoration of barrier function during wound healing process and infection with the common wound pathogen Staphylococcus aureus. We describe a novel approach for the measurement of P‐2 mRNA within individual skin cells using an amplified fluorescence in situ hybridization (FISH) technique. The unique aspect of this approach is simultaneous detection of P‐2 mRNA in combination with immune‐phenotyping for cell surface proteins using fluorochrome‐conjugated antibodies. We detected P‐2 transcript in both hematopoietic (CD45+) and non‐hematopoietic (CD45−) cutaneous cell populations, confirming the P‐2 expression in both professional and non‐professional phagocytes. Furthermore, we found an induction of P‐2 during wound healing. P‐2 overexpression resulted in a reduction of intracellular S. aureus, while infection of human wounds by this pathogen resulted in P‐2 suppression, revealing a novel mechanism by which S. aureus may escape cutaneous immunity to cause persistent wound infections. [ABSTRACT FROM AUTHOR]

Millions of people are infected each year by arboviruses (arthropod- borne viruses) such as chikungunya, dengue, and West Nile viruses, yet for reasons that are largely unknown, only a relatively small number of mosquito species are able to transmit arboviruses. Understanding the complex factors that determine vector competence could facilitate strategies for controlling arbovirus infections. Apoptosis is a potential antiviral defense response that has been shown to be important in other virus--host systems. However, apoptosis is rarely seen in arbovirus-infected mosquito cells, raising questions about its importance as an antiviral defense in mosquitoes. We tested the effect of stimulating apoptosis during arbovirus infection by infecting Aedes aegypti mosquitoes with a Sindbis virus (SINV) clone called MRE/Rpr, in which the MRE-16 strain of SINV was engineered to express the proapoptotic gene reaper from Drosophila. MRE/Rpr exhibited an impaired infection phenotype that included delayed midgut infection, delayed virus replication, and reduced virus accumulation in saliva. Nucleotide sequencing of the reaper insert in virus populations isolated from individual mosquitoes revealed evidence of rapid and strong selection against maintenance of Reaper expression inMRE/Rpr-infected mosquitoes. The impaired phenotype of MRE/Rpr, coupled with the observed negative selection against Reaper expression, indicates that apoptosis is a powerful defense against arbovirus infection in mosquitoes and suggests that arboviruses have evolved mechanisms to avoid stimulating apoptosis in mosquitoes that serve as vectors. [ABSTRACT FROM AUTHOR]

Perforin-2/mpeg-1, human skin, innate immunity, Staphylococcus epidermidis, gamma delta T cells, cytotoxicity Keywords: perforin-2/mpeg-1; human skin; innate immunity; Staphylococcus epidermidis; gamma delta T cells; cytotoxicity EN perforin-2/mpeg-1 human skin innate immunity Staphylococcus epidermidis gamma delta T cells cytotoxicity 1 2 2 08/13/21 20210810 NES 210810 In the original article, there was a mistake in Figure 1B as published. [Extracted from the article]

Bax and Bak, two functionally similar, pro-apoptotic proteins of the Bcl-2 family, are known as the gateway to apoptosis because of their requisite roles as effectors of mitochondrial outer membrane permeabilization (MOMP), a major step during mitochondria-dependent apoptosis. The mechanism of how cells turn Bax/Bak from inert molecules into fully active and lethal effectors had long been the focal point of a major debate centered around two competing, but not mutually exclusive, models: direct activation and indirect activation. After intensive research efforts for over two decades, it is now widely accepted that to initiate apoptosis, some of the BH3-only proteins, a subclass of the Bcl-2 family, directly engage Bax/Bak to trigger their conformational transformation and activation. However, a series of recent discoveries, using previously unavailable CRISPR-engineered cell systems, challenge the basic premise that undergirds the consensus and provide evidence for a novel and surprisingly simple model of Bax/Bak activation: the membrane (lipids)-mediated spontaneous model. This review will discuss the evidence, rationale, significance, and implications of this new model., Competing Interests: No competing interests were disclosed.No competing interests were disclosed.No competing interests were disclosed.Competing interests: Andreas Strasser is an employee of The Walter and Eliza Hall Institute. This Institute receives milestone payments and royalties from the sale of venetoclax. Andreas Strasser also has a collaboration with Servier for the development of MCXL01 inhibitors for cancer therapy., (Copyright: © 2020 Luo X et al.)

The anti-apoptotic Bcl-2 family protein Bcl-xL plays a critical role in cell survival by protecting the integrity of the mitochondrial outer membrane (MOM). The mechanism through which Bcl-xL is recruited to the MOM has not been fully discerned. The retromer is a conserved endosomal scaffold complex involved in membrane trafficking. Here we identify VPS35 and VPS26, two core components of the retromer, as novel regulators of Bcl-xL. We observed interactions and colocalization between Bcl-xL, VPS35, VPS26, and MICAL-L1, a protein involved in recycling endosome biogenesis that also interacts with the retromer. We also found that upon VPS35 depletion, levels of nonmitochondrial Bcl-xL were increased. In addition, retromer-depleted cells displayed more rapid Bax activation and apoptosis. These results suggest that the retromer regulates apoptosis by facilitating Bcl-xL's transport to the MOM. Importantly, our studies suggest a previously uncharacterized relationship between the machineries of cell death/survival and endosomal trafficking.

The mechanism of Bax/Bak activation remains a central question in mitochondria-dependent apoptotic signaling. While it is established that all proapoptotic Bcl-2 homology 3 (BH3)-only proteins bind and neutralize the anti-apoptotic Bcl-2 family proteins, how this neutralization leads to Bax/Bak activation has been actively debated. Here, genome editing was used to generate cells deficient for all eight proapoptotic BH3-only proteins (OctaKO) and those that lack the entire Bcl-2 family (Bcl-2 allKO). Although the OctaKO cells were resistant to most apoptotic stimuli tested, they underwent Bax/Bak-dependent and p53/Rb-independent apoptosis efficiently when both Bcl-xL and Mcl-1, two anti-apoptotic Bcl-2 proteins, were inactivated or eliminated. Strikingly, when expressed in the Bcl-2 allKO cells, both Bax and Bak spontaneously associated with the outer mitochondrial membrane (OMM) through their respective helix 9, and this association triggered their homo-oligomerization/activation. Together, these results strongly suggest that the OMM, not BH3-only proteins or p53/Rb, is the long-sought-after direct activator of Bax/Bak following BH3-only-mediated neutralization of anti-apoptotic Bcl-2 proteins., (© 2016 O'Neill et al.; Published by Cold Spring Harbor Laboratory Press.)