Candida albicans secreted protease Sap6 engages epithelial protease-activated receptors PAR2 and NLRP3

白色念珠菌分泌的蛋白酶 Sap6 与上皮蛋白酶激活受体 PAR2 和 NLRP3 结合

• 批准号: 10300121
• 负责人: Mira Edgerton
• 金额: $ 23.93万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10300121
• 关键词: Apoptosis; Aspartic Endopeptidases; Bacteria; Binding; Binding Sites; Blocking Antibodies; CASP1 gene; Calcium Signaling; Candida albicans; Candidiasis; Carbohydrates; Caspase; Catalytic Domain; Cell Surface Receptors; Cell Wall; Cell surface; Cells; Cellular Morphology; Data; Disseminated candidiasis; E-Cadherin; Electrical Resistance; Endocytosis; Epithelial; Epithelial Cells; FOS gene; Filament; Foundations; Fungal Proteins; Future; Goals; Human; Hyphae; IL17 gene; IL8 gene; Immunity; Immunocompromised Host; Inflammasome; Inflammation; Inflammatory Response; Integrin Binding; Integrins; Interleukin-1 beta; Knockout Mice; Measurement; Measures; Mediating; Morphology; Mucous Membrane; Mycoses; Neutrophil Infiltration; Opportunistic Infections; Oral; Pathogenicity; Pattern recognition receptor; Peptide Hydrolases; Peptides; Permeability; Phosphorylation; Production; Proteinase-Activated Receptors; Proteins; RGD (sequence); Receptor Signaling; Role; Signal Pathway; Signal Transduction; Site; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Surface; Symbiosis; Testing; Therapeutic; Virulence; Work; Yeasts; arm; cytokine; fungus; gut microbiota; in vivo Model; member; monocyte; mycobiome; novel; occludin; oral bacteria; oral cavity epithelium; p38 Mitogen Activated Protein Kinase; pathogenic fungus; receptor; receptor internalization; release of sequestered calcium ion into cytoplasm; treatment strategy

Candida albicans pathogenicity is associated with its morphological transformation from yeast to production of filamentous hyphae. Secretion of hyphal-specific proteins, particularly the aspartyl protease Sap6, is a key marker of its transition from commensalism to virulence. Sap6 contains both a proteinase domain and an RGDRGD integrin-binding motif adjacent to its catalytic domain. We found that rSap6 added to primary oral epithelial cells (OECs) elicited IL1β secretion typical of NLRP3 signaling but also produced IL-8. Sap6 added to OECs induced phosphorylation of p38 and IL-8 production through the protease- activated receptor (PAR2). PAR2 signaling was lost upon heat inactivation of Sap6, while NLRP signaling was reduced upon deletion of Sap6 RGD-binding sites, leading to our hypothesis that both Sap6 proteolytic and RGD-mediated mechanisms are involved in sensing levels of this fungal secreted protein by OECs. We will test this hypothesis by identifying the proteolytic mechanism for Sap6 activated PAR2 MAPKinase signaling in OECs in Aim1, and assessing the contribution of the Sap6 integrin-binding RGD motif in NLRP3 inflammasome activation and apoptosis in Aim 2. This work will establish PAR2 and integrin/NLRP3 as cell surface receptors for C. albicans Sap6 and define their role in modulating host inflammatory responses to fungal proteins in OECs. The long-range goal of this project will be to activate/deactivate PAR2 receptors or NLRP3 signaling as a means of modulating host inflammation as an avenue for treatment of candidiasis.

白色念珠菌的致病性与其从酵母菌到 丝状菌丝的产生。菌丝特异性蛋白质的分泌，特别是 蛋白酶Sap 6是其从嗜热性向毒力转变的关键标志。sap 6包含 蛋白酶结构域和邻近其催化结构域的RGDRGD整联蛋白结合基序。 我们发现，rSap 6加入到原代口腔上皮细胞（OECs）中可引起典型的IL 1 β分泌， NLRP 3信号传导，但也产生IL-8。向OECs中加入Sap 6诱导p38磷酸化， 通过蛋白酶激活受体（PAR 2）产生IL-8。PAR 2信号传导在加热时丢失 Sap 6的失活，而NLRP信号传导在Sap 6 RGD结合位点缺失后减少， 这导致我们假设Sap 6蛋白水解和RGD介导的机制都参与了 通过OECs检测这种真菌分泌蛋白的水平。我们将通过识别 Aim 1中OEC中Sap 6激活PAR 2 MAP激酶信号传导的蛋白水解机制，以及 评估NLRP 3炎性体中Sap 6整联蛋白结合RGD基序的贡献 Aim 2中的活化和凋亡。这项工作将建立PAR 2和整合素/NLRP 3作为细胞表面 C.白色念珠菌Sap 6，并确定其在调节宿主炎症反应， OECs中的真菌蛋白。本项目的长期目标是激活/停用PAR 2 受体或NLRP 3信号传导作为调节宿主炎症的手段， 治疗念珠菌病。