Project No.2235

Primary Supervisor

Dr Anthony Lewis – University of Portsmouth

Co-Supervisor(s)

Dr Ruth Murrell-Lagnado – University of Sussex

Prof. John Spencer – University of Sussex

Summary

Rationale: Opportunistic fungal pathogens are a significant threat to human health and global food security.

The effectiveness of current fungicides to treat human infections and agricultural blights is in rapid decline due to evolving fungal resistance, and is complicated by biological similarities between fungi and their hosts. TOK ion channels are unique plasma membrane potassium channels found only in fungi, and no structurally or functionally equivalent protein exists in humans, animals or plants. We have recently

cloned and started characterisation of the basal biophysical properties of TOK channels cloned from multiple species of human and plant fungal pathogens. Given their functional attributes, data from the Lewis lab (UoP), and other published work, indicates TOK channels could be novel strategic targets for future antimicrobial compounds. We hypothesise that perpetual TOK activation will lead to changes in virulence-associated cell morphology and cell death. Aims and Approach: Here, we will be to continue the detailed structure-function characterisation of TOK channels using two-electrode voltage-clamp (TEVC) and patch clamp electrophysiology (whole-cell and single-channel), focussing on their activation by environmental and mechanical stimuli. This profiling will help us enable detail the functional role of TOK channels in fungal cell physiology, morphology and virulence using the model yeast, Candida albicans. The final aim will be to assay for potent chemical interactors of the TOK channel that demonstrate good efficacy in established fungal cell death and morphology assays. This will involve the creation of stable cells lines and implementation of fluorescence-based flux screening assays as well as orthogonal assays using QPatch technology and screening of a 2K LifeChem ion channel CUTE compound library. Impact: The unique fungal specific nature of TOK channels coupled with their potential role in fungal cell death and morphology, through promotion of ion dyshomeostasis, makes them viable targets for future fungicidal compounds to combat deleterious human, animal and agricultural mycoses.