Media type: E-Article Title: Quantitative Proteomic Profiling of Cryptococcus neoformans Contributor: Ball, Brianna; Geddes‐McAlister, Jennifer Published: Wiley, 2019 Published in: Current Protocols in Microbiology, 55 (2019) 1 Language: English DOI: 10.1002/cpmc.94 ISSN: 1934-8525; 1934-8533 Keywords: Virology ; Microbiology ; Parasitology Origination: Footnote: Description: <jats:title>Abstract</jats:title><jats:p><jats:italic>Cryptococcus neoformans</jats:italic> is an opportunistic human fungal pathogen commonly associated with infection in immunocompromised individuals (e.g., patients with HIV/AIDS). Important virulence determinants include the production of a polysaccharide capsule, melanin, and extracellular enzymes, as well as the ability to grow at 37°C. <jats:italic>C. neoformans</jats:italic> controls a plethora of host defense and evasion mechanisms to survive during infection and to proliferate within the host, causing meningoencephalitis and death. Traditionally, characterization of <jats:italic>C. neoformans</jats:italic> under different environmental conditions and stresses has relied on genetic and phenotypic analyses, as well as biochemical assays. However, advances in mass spectrometry instrumentation, sample preparation protocols, and bioinformatic tools and databases promote comprehensive profiling of fungal cellular processes, secretion or protein release into the extracellular environment, and vesicle contents. Moreover, proteomics provides insight into regulatory mechanisms influencing signal transduction cascades and protein complexes or networks through profiling of post‐translational modifications and protein–protein interactions. Given the medical impact of <jats:italic>C. neoformans</jats:italic> infections and the recent emergence of antifungal‐resistant strains, defining proteins produced in response to unique environments provides an opportunity to uncover antivirulence strategies and alternative therapeutic options to combat infection. Here, we describe culturing and sample preparation of <jats:italic>C. neoformans</jats:italic> and outline protocols for comprehensively profiling changes in protein abundance within the cellular proteome and secretome. © 2019 by John Wiley & Sons, Inc.</jats:p><jats:p><jats:bold>Basic Protocol 1</jats:bold>: Growth and sample preparation of <jats:italic>Cryptococcus neoformans</jats:italic></jats:p><jats:p><jats:bold>Basic Protocol 2</jats:bold>: Protein extraction from supernatant</jats:p><jats:p><jats:bold>Basic Protocol 3</jats:bold>: Protein extraction from cell pellet</jats:p><jats:p><jats:bold>Basic Protocol 4</jats:bold>: Proteomic profiling and bioinformatics</jats:p>