Cryo-Electron Tomography of Candida glabrata Plasma Membrane Proteins.

Cristina Jiménez-Ortigosa,Muyuan Chen,Steven J Ludtke,Jennifer Jiang,David S Perlin,Wei Dai,Kelley R Healey,Paul Castellano,Nikpreet Boparai,Xuyuan Kuang

doi:10.3390/jof7020120

Abstract

Echinocandin drugs have become a front-line therapy against Candida spp. infections due to the increased incidence of infections by species with elevated azole resistance, such as Candida glabrata. Echinocandins target the fungal-specific enzyme ß-(1,3)-glucan synthase (GS), which is located in the plasma membrane and catalyzes the biosynthesis of ß-(1,3)-glucan, the major component of the fungal cell wall. However, resistance to echinocandin drugs, which results from hotspot mutations in the catalytic subunits of GS, is an emerging problem. Little structural information on GS is currently available because, thus far, the GS enzyme complex has resisted homogenous purification, limiting our understanding of GS as a major biosynthetic apparatus for cell wall assembly and an important therapeutic drug target. Here, by applying cryo-electron tomography (cryo-ET) and subtomogram analysis, we provide a preliminary structure of the putative C. glabrata GS complex as clusters of hexamers, each subunit with two notable cytosolic domains, the N-terminal and central catalytic domains. This study lays the foundation for structural and functional studies of this elusive protein complex, which will provide insight into fungal cell wall synthesis and the development of more efficacious antifungal therapeutics.

Highlights

Invasive mycoses have become a significant threat to public health, affecting over a billion people globally and causing over 1.1 million deaths per year
We examined over 600 tomograms and found that 8–10% contained clusters of a ring-like structure with an approximate diameter of 170 Å
We present the first study describing an initial structure of the putative glucan synthase (GS) complex from the pathogen Candida glabrata using Cryo-electron tomography (cryoET), a powerful technique for 3D visualization of cellular structures, especially membrane proteins in their native state

Summary

Introduction

Invasive mycoses have become a significant threat to public health, affecting over a billion people globally and causing over 1.1 million deaths per year. Species from the genera Aspergillus, Candida and Cryptococcus are the most common human pathogenic fungi accounting for a wide variety of invasive and superficial fungal infections. Underlying health conditions such as asthma, acquired immunodeficiency syndrome (AIDS), diabetes, cancer, organ transplantation, and use of corticosteroid therapy are important risk factors for invasive disease. And appropriate antifungal treatment is crucial for the successful outcome of invasive fungal infections [1,2,3,4]. The fungal cell wall is an essential dynamic structure that undergoes extensive remodeling necessary for growth, survival, fungal morphogenesis and pathogenesis, and protection against osmotic and mechanical stresses. Since the fungal cell wall does not have

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