Unraveling the complexities of signaling and secretion within the fungal pathogen, Cryptococcus neoformans

Summary

Research opportunities are available to decipher the complexities of virulence-related signalling and secretion pathways in pathogenic fungi, and to investigate the mechanisms of action of novel antifungal compounds, using molecular and cell biology techniques.

Supervisor(s)

Dr Julianne Djordjevic, Professor Tania Sorrell

Research Location

Westmead - Sydney Emerging Infections and Biosecurity Institute

Program Type

Masters/PHD

Synopsis

Unraveling the complexities of signaling and secretion within the fungal pathogen, Cryptococcus neoformansCryptococcus neoformans is a common cause of invasive fungal infection and is responsible for an estimated one million cases of meningitis and 675,000 deaths per year in HIV-infected patients alone. In light of the scarcity of antifungal drug classes, the emergence of resistant fungi and the unfavourable toxicity/bioavailability profiles of key marketed antifungals, the search for new drug targets is of considerable importance. Cryptococcus neoformans is also an excellent fungal pathogenicity model, as its sequenced genome is highly amenable to manipulation and robust animal models of infection are available.

Project 1- Identifying secretion mechanisms in pathogenic fungi
Fungal secretion pathways are complex and crucial for maintaining fungal fitness, yeast turgor pressure (by ensuring production of a robust cell wall) and pathogenicity by allowing the externalization of an array of virulence factors including the common fungal invasion-promoting enzyme phospholipase B1, polysaccaharide components of the capsular (assists in the evasion of a human immune response) and the melanin-producing enzyme laccase. Externalized melanin protects against oxidative stress that is associated with host immune defense mechanisms. Our research is focused on identifying yeast secretion pathways and their components, and determining the role of the gene, SEC14, in phospholipase B1 secretion and pathogenicity.

Project 2- Delineating Plc1-dependent signaling pathways involved in fungal pathogenicity
The phenotypic expression of many known fungal pathogenicity determinants is regulated by a number of signal transduction pathways. However, many "gaps" in these signaling networks remain to be elucidated. Our recent molecular investigations in C. neoformans identified a gene encoding a phospholipase C (Plc1) which is essential for the phenotypic expression a number of virulence determinants, including secretion of phospholipase B1. Epistasis-based research projects are available to investigate the role of Plc1 in virulence-related signaling. Expected outcomes of Project 1 and 2 are the identification of potential molecular targets for anti-fungal drug development.

Project 3 - Identifying the site of action of Miltefosine-related antifungal compounds.
Opportunities are also available to determine the site of action of several antifungal compounds synthesized by the School of Chemistry. These compounds are based on the drug, Miltefosine, which although is in clinical use to treat parasitic infections, also displays antifungal properties in vitro and in vivo.

Techniques
Techniques include targeted gene disruption/silencing using overlap PCR and biolistic transformation, semi-quantitative real-time PCR to compare mRNA expression levels, southern and western blotting, cloning/creation of fungal cDNA libraries, mutagenesis, transcriptomics, proteomics, metabolomics, assessing virulence phenotypes and cell wall robustness in vitro, and in vivo using animal and cell infection models. Constructing fluorescently-tagged molecules and tracking their intracellular location/secretion using fluorescence microscopy. Macrophage cell culture, susceptibility assays, drug toxicity testing.

Keywords

Cryptococcus neoformans/cryptococcosis, HIV/AIDS, Virulence, Phospholipase B, Secretion pathways, SEC14, Fungal cell wall integrity, Host-microbe interactions, Drug targets, Miltefosine, Infectious diseases, Pathogenic fungi, macrophages

Opportunity ID

The opportunity ID for this research opportunity is: 1149

Other opportunities with Dr Julianne Djordjevic

• Elucidating secretory pathways and mechanisms essential for establishing cell wall integrity in pathogenic fungi
• Unraveling the complexities of signaling and secretion within the fungal pathogen, Cryptococcus neoformans

Other opportunities with Professor Tania Sorrell

• Unraveling the complexities of signaling and secretion within the fungal pathogen, Cryptococcus neoformans