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

Award Number: R01AI167692
ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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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.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2023 ( Subtotal = $382,675 )
2023	2023	CLEMSON UNIVERSITY	201 SIKES HALL	CLEMSON	SC	29634	PICKENS	USA	Allergy and Infectious Diseases Research	000	2	7/25/2023	NON-COMPETING CONTINUATION	$382,675
														Subtotal = $382,675

Issue Date FY: 2022 ( Subtotal = $356,944 )
2022	2022	CLEMSON UNIVERSITY	201 SIKES HALL	CLEMSON	SC	29634	PICKENS	USA	Allergy and Infectious Diseases Research	000	1	9/21/2022	NEW	$356,944
														Subtotal = $356,944

Grand Total All Awards = $739,619

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

Award Number: R01AI167692

ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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