Cryptococcus neoformans Iron-Sulfur Protein Biogenesis Machinery Is a Novel Layer of Protection against Cu Stress.

Sarela Garcia-Santamarina,Roland Lill,Dennis J Thiele,Richard A Festa,Marta A Uzarska,Michael Lorenz

doi:10.1128/mbio.01742-17

Abstract

ABSTRACTCopper (Cu) ions serve as catalytic cofactors to drive key biochemical processes, and yet Cu levels that exceed cellular homeostatic control capacity are toxic. The underlying mechanisms for Cu toxicity are poorly understood. During pulmonary infection by the fungal pathogen Cryptococcus neoformans, host alveolar macrophages compartmentalize Cu to the phagosome, and the ability to detoxify Cu is critical for its survival and virulence. Here, we report that iron-sulfur (Fe-S) clusters are critical targets of Cu toxicity in both Saccharomyces cerevisiae and C. neoformans in a manner that depends on the accessibility of Cu to the Fe-S cofactor. To respond to this Cu-dependent Fe-S stress, C. neoformans induces the transcription of mitochondrial ABC transporter Atm1, which functions in cytosolic-nuclear Fe-S protein biogenesis in response to Cu and in a manner dependent on the Cu metalloregulatory transcription factor Cuf1. As Atm1 functions in exporting an Fe-S precursor from the mitochondrial matrix to the cytosol, C. neoformans cells depleted for Atm1 are sensitive to Cu even while the Cu-detoxifying metallothionein proteins are highly expressed. We provide evidence for a previously unrecognized microbial defense mechanism to deal with Cu toxicity, and we highlight the importance for C. neoformans of having several distinct mechanisms for coping with Cu toxicity which together could contribute to the success of this microbe as an opportunistic human fungal pathogen.

Highlights

Copper (Cu) ions serve as catalytic cofactors to drive key biochemical processes, and yet Cu levels that exceed cellular homeostatic control capacity are toxic
One gene activated during Cu stress corresponds to CNAG_04358, which codes for a homologue of the previously characterized S. cerevisiae mitochondrial inner membrane ATPbinding cassette (ABC) transporter Atm1
To further address whether C. neoformans ATM1 belongs to the Cuf1 regulon, Cuf1 occupancy on the ATM1 promoter was analyzed by chromatin immunoprecipitation-PCR (ChIP-PCR) in wild-type cells that were treated either with Cu or with the Cu chelator bathocuproine disulfonic acid (BCS) (Fig. 1B)

Summary

Introduction

Copper (Cu) ions serve as catalytic cofactors to drive key biochemical processes, and yet Cu levels that exceed cellular homeostatic control capacity are toxic. During pulmonary infection by the fungal pathogen Cryptococcus neoformans, host alveolar macrophages compartmentalize Cu to the phagosome, and the ability to detoxify Cu is critical for its survival and virulence. When present at high concentrations, Cu is toxic In this regard, C. neoformans Cu metalloregulatory transcription factor Cuf has been shown to be required for virulence, as it regulates the expression of genes required for both the acquisition and detoxification of Cu [17, 18]. Defects in Cuϩ exporters or in metallothionein expression sensitize both bacterial and fungal pathogens to macrophage killing mechanisms that are dependent on the toxicity of Cu [20,21,22,23]. Additional work performed in vitro demonstrated that Cuϩ destabilizes Fe-S clusters from the bacterial SufU protein [27], the major scaffold used by the sulfur assimilation (SUF) system for Fe-S cluster assembly and transfer to target proteins, and from mammalian ISCA1/2 and GLRX5, mitochondrial proteins involved in Fe-S cluster trafficking [28]

Methods

Results

Conclusion