Tolerance to alkaline ambient pH in Aspergillus nidulans depends on the activity of ENA proteins

Ane Markina-Iñarrairaegui,Unai Ugalde,Oier Etxebeste,Eduardo A Espeso,Anja Spielvogel

doi:10.1038/s41598-020-71297-z

Abstract

Tolerance of microorganisms to abiotic stress is enabled by regulatory mechanisms that coordinate the expression and activity of resistance genes. Alkalinity and high salt concentrations are major environmental physicochemical stresses. Here, we analyzed the roles of sodium-extrusion family (ENA) transporters EnaA, EnaB and EnaC in the response to these stress conditions in the filamentous fungus Aspergillus nidulans. While EnaC has a minor role, EnaB is a key element for tolerance to Na+ and Li+ toxicity. Adaptation to alkaline pH requires the concerted action of EnaB with EnaA. Accordingly, expression of enaA and enaB was induced by Na+, Li+ and pH 8. These expression patterns are altered in a sltAΔ background and completely inhibited in a mutant expressing non-functional PacC protein (palH72). However, a constitutively active PacC form was not sufficient to restore maximum enaA expression. In agreement with their predicted role as membrane ATPases, EnaA localized to the plasma membrane while EnaB accumulated at structures resembling the endoplasmic reticulum. Overall, results suggest different PacC- and SltA-dependent roles for EnaB in pH and salt homeostasis, acting in coordination with EnaA at pH 8 but independently under salt stress.

Highlights

Ion homeostasis is an essential biochemical process for cell life influencing a wide range of cellular functions from osmotic regulation to enzyme activity
A more detailed analysis of A. nidulans extrusion family (ENA) protein sequences compared to ScENA1p showed the presence of several small motifs characteristic of this family of transporters, such as the Actuator domain (TGES183), the Mg2+ binding motif (DGVND761), and sequences belonging to the catalytic site of P-type ATPases: the signature sequences DKTGT393, TGD675 and D PPR652 and residues of the nucleotide binding domain (F537, K542, K561)[15,27]
In this work we investigated, through a combination of genetics and cell biology techniques, the roles of the three putative sodium ATPases of the ENA family identified in A. nidulans[16,19,38]

Summary

Introduction

Ion homeostasis is an essential biochemical process for cell life influencing a wide range of cellular functions from osmotic regulation to enzyme activity. The activity of plasma-membrane transporters generates an electrochemical gradient across the membrane which stores energy that can be used by other transport mechanisms acting as antiporters and/or symporters In this respect, the transmembrane electrochemical proton gradient has a major influence on the transmembrane potential and on the functionality of ion transport across the plasma membrane in both directions[6]. A change in the extracellular pH, either towards acidification or to alkalinization, is an important stimulus to a cell or organism This might cause alterations in the electrochemical gradient and its maintenance, with direct impact on the bidirectional transport of solutes and other compounds. In S. cerevisiae intracellular low sodium maintenance relies on the Nha1p N a+/ H+ antiporter which is able to extrude N a+, K+, and Li+11 These antiporters depend on the transmembrane ∆pH, being less functional when the external pH is higher than the cytoplasmic pH, resulting in a decreased pH gradient. P-type Ena ATPases mediate Na+/K+ transport under alkaline c onditions[5]

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Results

Conclusion