The role of septins in the adaptation of Cryptococcus neoformans to host temperature in HIV-based cryptococcosis

脓毒症在 HIV 隐球菌病中新型隐球菌适应宿主温度中的作用

• 批准号: 10619216
• 负责人: Lukasz Kozubowski
• 金额: $ 35.69万
• 依托单位: CLEMSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-21 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10619216
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Address; Alleles; Animal Model; Animals; Aspergillus nidulans; Binding; Blood Circulation; Cell Membrane Permeability; Cell Wall; Cell membrane; Cells; Central Nervous System Infections; Cessation of life; Complex; Critical Pathways; Cryptococcosis; Cryptococcus; Cryptococcus neoformans; Cues; Cytokinesis; Data; Drug Targeting; Engineering; Exhibits; Exposure to; Fatal Outcome; Filament; Fluconazole; Fluorescence Microscopy; Foundations; Genes; Genetic; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; HIV; High temperature of physical object; Homeostasis; Human; Hypersensitivity; Immunocompromised Host; In Vitro; Infection; Inhalation; Inositol Phosphates; Knowledge; Life; Light; Link; Lipid Bilayers; Lipids; Lung; Maintenance; Mediator of activation protein; Membrane; Membrane Fluidity; Membrane Lipids; Meningoencephalitis; Modeling; Monomeric GTP-Binding Proteins; Mus; Outcome; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacology; Phenotype; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositols; Phospholipase C; Population; Propidium Diiodide; Protein Kinase C; Proteins; Public Health; Publishing; Recombinants; Research; Role; Signal Pathway; Signal Transduction; Sodium Dodecyl Sulfate; Sphingolipids; Sterols; Stress; Stress Response Signaling; Temperature; Temperature Sense; Testing; Time; Virulence; base; biophysical properties; experimental study; fluidity; genetic approach; improved; inhibitor; innovation; lipid metabolism; mortality; mutant; novel; pathogenic fungus; predictive modeling; prevent; protein complex; reconstitution; recruit; response; side effect; temperature sensitive mutant; thermozymocidin; treatment strategy

PROJECT SUMMARY The long-term goal of this proposal is to uncover mechanisms of pathogenicity of Cryptococcus neoformans (Cn) focusing on how Cn adapts to host temperature. Cn is a fungal pathogen that, upon entering the lung and disseminating through the bloodstream, causes a life-threatening meningo-encephalitis primarily in HIV/AIDS patients. Current anti-cryptococcal therapies can have devastating side effects. Thus, new treatment strategies are warranted to eliminate mortality associated with cryptococcosis. The objective of this proposal is to determine how the temperature is sensed by Cn and transmitted to downstream effector pathways critical for pathogenicity. The central hypothesis is that filament-forming GTP-ases called septins provide an essential hub protein complex which links temperature sensing to a compensatory signaling response and plasma membrane (PM) homeostasis. It is proposed that exposure to host temperature results in increased fluidity and curvature within the PM and the elevation of PM-associated phosphatidylinositol 4,5-bisphosphate (PIP2) leading to enrichment of septins at the PM based on PIP2 binding, which has two-fold role in Cn adaptation to host temperature: it maintains PM homeostasis and facilitates stress signaling via phospholipase C (PLC), and protein kinase C (PKC) pathways. In support of this hypothesis, experiments with recombinant septins or studies employing model organisms suggest that septins recognize membrane curvature via their propensity to bind PIP2. Importantly, recombinant septins prevent temperature-induced changes in lipid bilayer composition. Preliminary data supporting this application are: 1) Cn PIP2 levels increase at 37°C. 2) Septins associate with the PM when Cn is shifted to 37°C. 3) Septin-deficient mutants exhibit increased PM permeability and sensitivity to drugs that perturb PM. Published and preliminary data also demonstrate phenotypic similarity between septin-deficient and PLC signaling-defective mutants. The central hypothesis will be tested by pursuing two aims. Aim1: Dynamics of septin assembly at the PM will be defined with TIRF microscopy. Genetic and pharmacological approaches will determine whether septin complexes are enriched at the PM based on increased levels of PIP2. An innovative light switchable allele of Cdc42 (GTPase involved in septin assembly) will help to establish effectors down-stream of Cdc42 acting in septin complex formation. A septin mutant lacking the amphiphatic helix (AH) will be utilized to test the role of AH in recruiting septin complex to the PM. Aim2: An impact of septin complexes on PM lipid composition, PM biophysical properties, stress response signaling dependent on PIP2, and pathogenesis in animal infection model will be determined. The proposed research is significant because it will elucidate a novel mechanism through which septins contribute to sensing high temperature and regulating PM-dependent stress response signaling pathways crucial for the pathogenesis of Cn. Outcomes will be better understanding of how Cn adapts to host temperature and a new foundation upon which to develop improved anti-cryptococcal therapies in HIV/AIDS patients.

项目总结