Monitoring Glycolysis and Respiration Highlights Metabolic Inflexibility of Cryptococcus neoformans

Pathogens

Desmarini Desmarini¹  JulianneT. Djordjevic¹  Cecilia Li¹  TaniaC. Sorrell¹  Sophie Lev¹  David Liuwantara²  WayneJ. Hawthorne³ 

[1] Centre for Infectious Diseases and Microbiology, The Westmead Institute for Medical Research, Westmead, NSW 2145, Australia;Centre for Transplant and Renal Research, The Westmead Institute for Medical Research, Westmead, NSW 2145, Australia;Sydney Medical School—Westmead, The University of Sydney, Westmead, NSW 2145, Australia;

关键词: Cryptococcus neoformans;    seahorse analyzer;    ECAR;    OCR;    metabolic flux;    glycolysis;   

DOI  :  10.3390/pathogens9090684

来源: DOAJ

【 摘 要 】

Cryptococcus neoformans is a human fungal pathogen that adapts its metabolism to cope with limited oxygen availability, nutrient deprivation and host phagocytes. To gain insight into cryptococcal metabolism, we optimized a protocol for the Seahorse Analyzer, which measures extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) as indications of glycolytic and respiratory activities. In doing so we achieved effective immobilization of encapsulated cryptococci, established Rotenone/Antimycin A and 2-deoxyglucose as effective inhibitors of mitochondrial respiration and glycolysis, respectively, and optimized a microscopy-based method of data normalization. We applied the protocol to monitor metabolic changes in the pathogen alone and in co-culture with human blood-derived monocytes. We also compared metabolic flux in wild-type C. neoformans, its isogenic 5-PP-IP₅/IP₇-deficient metabolic mutant kcs1∆, the sister species of C. neoformans, Cryptococcus deuterogattii/VGII, and two other yeasts, Saccharomyces cerevisiae and Candida albicans. Our findings show that in contrast to monocytes and C. albicans, glycolysis and respiration are tightly coupled in C. neoformans and C. deuterogattii, as no compensatory increase in glycolysis occurred following inhibition of respiration. We also demonstrate that kcs1∆ has reduced metabolic activity that correlates with reduced mitochondrial function. Metabolic inflexibility in C. neoformans is therefore consistent with its obligate aerobe status and coincides with phagocyte tolerance of ingested cryptococcal cells.

【 授权许可】

Unknown