Proteomics reveals distinct mechanisms regulating the release of cytokines and alarmins during pyroptosis.

A major pathway for proinflammatory protein release by macrophages is inflammasome-mediated pyroptotic cell death. As conventional secretion, unconventional secretion, and cell death are executed simultaneously, however, the cellular mechanisms regulating this complex paracrine program remain incompletely understood. Here, we devise a quantitative proteomics strategy to define the cellular exit route for each protein by pharmacological and genetic dissection of cellular checkpoints regulating protein release. We report the release of hundreds of proteins during pyroptosis, predominantly due to cell lysis. They comprise constitutively expressed and transcriptionally induced proteins derived from the cytoplasm and specific intracellular organelles. Many low-molecular-weight proteins including the cytokine interleukin-1ß, alarmins, and lysosomal-cargo proteins exit cells in the absence of cell lysis. Cytokines and alarmins are released in an endoplasmic reticulum (ER)-Golgi-dependent manner as free proteins rather than by extracellular vesicles. Our work provides an experimental framework for the dissection of cellular exit pathways and a resource for pyroptotic protein release.

Candidalysin Is a Potent Trigger of Alarmin and Antimicrobial Peptide Release in Epithelial Cells.

Host released alarmins and antimicrobial peptides (AMPs) are highly effective as antifungal agents and inducers. Whilst some are expressed constitutively at mucosal tissues, the primary site of many infections, others are elicited in response to pathogens. In the context of Candida albicans, the fungal factors inducing the release of these innate immune molecules are poorly defined. Herein, we identify candidalysin as a potent trigger of several key alarmins and AMPs known to possess potent anti-Candida functions. We also find extracellular ATP to be an important activator of candidalysin-induced epithelial signalling responses, namely epidermal growth factor receptor (EGFR) and MAPK signalling, which mediate downstream innate immunity during oral epithelial infection. The data provide novel mechanistic insight into the induction of multiple key alarmins and AMPs, important for antifungal defences against C. albicans.

Inflammasome as a promising therapeutic target for cancer.

Inflammasomes are the major mechanistic complexes that include members of the NOD-like receptor (NLRs) or AIM2-like receptors (ALRs) families, which are affiliated with the innate immune system. Once NLRs or ALRs are activated by pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs), the caspase-1 or -11 is activated by binding with NLRs or ALRs via its own unique cytosolic domains. As a result, caspase-1 or -11 enhances the production of IL-1ß and IL-18, which results in inflammation via the recruitment of immune cells, such as macrophages, and the promotion of programmed cell death mechanisms such as pyroptosis. In addition, the consistent cascades of inflammasomes would precede both minor and severe autoimmune diseases and cancers. The clinical relevance of inflammasomes in multiple forms of cancer highlights their therapeutic promise as molecular targets. To closely analyze the physiological roles of inflammasomes in cancers, here, we describe the fundamental knowledge regarding the current issues of inflammasomes in relevant cancers, and discuss possible therapeutic values in targeting these inflammasomes for the prevention and treatment of cancer.

Insights into the cardioprotective properties of n-3 PUFAs against ischemic heart disease via modulation of the innate immune system.

Ischemic heart disease (IHD) is a major cause of cardiovascular morbidity and mortality worldwide, which is characterized by an imbalance between cardiac oxygen supply and demand predominantly due to obstruction of coronary arteries. Activation of the innate immune system and the consequent inflammatory response plays a role in the pathogenesis of IHD. Where an excessive inflammatory response may contribute to adverse cardiac remodeling and fibrosis, making inflammation an important therapeutic target for improving outcomes of IHD. While there are many discrepancies in the literature, evidence from both bench and clinical research demonstrate important effects of n-3 polyunsaturated fatty acids (n-3 PUFA), eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA), toward IHD. N-3 PUFAs, and their metabolites, have been demonstrated to modulate various components of the immune system, including regulation of chemokines and cytokines, leukocyte chemotaxis and inflammasome formation. In this article, we provide an overview of the role the innate immune system has in IHD and focus on the immunomodulatory effects of n-3 PUFAs and their biologically active metabolites.

The oomycete microbe-associated molecular pattern Pep-13 triggers SERK3/BAK1-independent plant immunity.

KEY MESSAGE: Oomycetes MAMP Pep-13 can trigger SERK3/BAK1-independent PTI. Silencing of SERK3/BAK1 in solanaceous plants resulted in reduced expression of brassinosteroid marker genes and enhanced PTI transcriptional responses to Pep-13 treatment. To prevent disease, pattern recognition receptors (PRRs) are responsible for detecting microbe-associated molecular patterns (MAMPs) to switch on plant innate immunity. SOMATIC EMBROYOGENESIS KINASE 3 (SERK3)/BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE 1 (BAK1) is a well-characterized receptor-like kinase (RLK) that serves as a pivotal co-receptor with PRRs to activate immunity following recognition of MAMPs including flg22, EF-Tu, INF1 and XEG1. However, the requirement for SERK3/BAK1 in many pattern-triggered immune (PTI) signaling pathways is not yet known. Pep-13 is an oomycete MAMP that consists of a highly conserved motif (an oligopeptide of 13 amino acids) shared in Phytophthora transglutaminases. Quantitative RT-PCR analysis reveals that the transcripts of three PTI marker genes (WRKY7, WRKY8 and ACRE31) rapidly accumulate in response to three different MAMPs: flg22, chitin and Pep-13. Whereas silencing of SERK3/BAK1 in Nicotiana benthamiana or potato compromised transcript accumulation in response to flg22, it did not attenuate WRKY7, WRKY8 and ACRE31 up-regulation in response to chitin or Pep-13. This indicates that Pep-13 triggers immunity in a SERK3/BAK1-independent manner, similar to chitin. Surprisingly, silencing of SERK3/BAK1 led to significantly increased accumulation of PTI marker gene transcripts following Pep-13 or chitin treatment, compared to controls. This was accompanied by reduced expression of brassinosteroid (BR) marker genes StSTDH, StEXP8 and StCAB50 and StCHL1, which is a negative regulator of PTI, supporting previous reports that SERK3/BAK1-dependent BR signaling attenuates plant immunity. We provide Pep-13 as an alternative to chitin as a trigger of SERK3/BAK1-independent immunity.

AIF-1 and RNASET2 Play Complementary Roles in the Innate Immune Response of Medicinal Leech.

Recent studies demonstrated that allograft inflammatory factor-1 (AIF-1) and RNASET2 act as chemoattractants for macrophages and modulate the inflammatory processes in both vertebrates and invertebrates. The expression of these proteins significantly increases after bacterial infection; however, the mechanisms by which they regulate the innate immune response are still poorly defined. Here, we evaluate the effect of bacterial lipopolysaccharide injection on the expression pattern of these genes and the interrelation between them during innate immune response in the medicinal leech, an invertebrate model with a simple anatomy and a marked similarity with vertebrates in inflammatory processes. Collectively, prokaryotic-eukaryotic co-cultures and in vivo infection assays suggest that RNASET2 and AIF-1 play a crucial role in orchestrating a functional cross-talk between granulocytes and macrophages in leeches, resulting in the activation of an effective response against pathogen infection. RNASET2, firstly released by granulocytes, likely plays an early antibacterial role. Subsequently, AIF-1+ RNASET2-recruited macrophages further recruit other macrophages to potentiate the antibacterial inflammatory response. These experimental data are in keeping with the notion of RNA-SET2 acting as an alarmin-like molecule whose role is to locally transmit a "danger" signal (such as a bacterial infection) to the innate immune system in order to trigger an appropriate host response.