Clotrimazole-Induced Oxidative Stress Triggers Novel Yeast Pkc1-Independent Cell Wall Integrity MAPK Pathway Circuitry

Abstract

Azoles are one of the most widely used drugs to treat fungal infections. To further understand the fungal response to azoles, we analyzed the MAPK circuitry of the model yeast Saccharomyces cerevisiae that operates under treatment with these antifungals. Imidazoles, and particularly clotrimazole, trigger deeper changes in MAPK phosphorylation than triazoles, involving a reduction in signaling through the mating pathway and the activation of the MAPKs Hog1 and Slt2 from the High-Osmolarity Glycerol (HOG) and the Cell Wall Integrity (CWI) pathways, respectively. Clotrimazole treatment leads to actin aggregation, mitochondrial alteration, and oxidative stress, which is essential not only for the activation of both MAPKs, but also for the appearance of a low-mobility form of Slt2 caused by additional phosphorylation to that occurring at the conserved TEY activation motif. Clotrimazole-induced ROS production and Slt2 phosphorylation are linked to Tpk3-mediated PKA activity. Resistance to clotrimazole depends on HOG and CWI-pathway-mediated stress responses. However, Pkc1 and other proteins acting upstream in the pathway are not critical for the activation of the Slt2 MAPK module, suggesting a novel rewiring of signaling through the CWI pathway. We further show that the strong impact of azole treatment on MAPK signaling is conserved in other yeast species.

Involvement of the High-Osmolarity Glycerol Pathway of Saccharomyces Cerevisiae in Protection against Copper Toxicity

“…To ascertain the association of oxidative stress with HOG signaling following copper stimulation, the ROS scavenger N-acetylcysteine (NAC) was employed in this study [ 34 ]. After adding NAC, copper’s inhibitory effect on yeast is significantly weakened ( Figure 4 A).…”

Organophosphorus Azoles Incorporating a Tetra-, Penta-, and Hexacoordinated Phosphorus Atom: NMR Spectroscopy and Quantum Chemistry

“…Azoles are the structural core of the most popular drugs used for the treatment of various diseases. Interest in organophosphorus compounds, including those containing azolyl moieties, increases every year due to their high and unusual (specific) biological activity [ 69 , 70 , 71 , 72 , 73 , 74 , 75 , 76 , 77 , 78 , 79 , 80 ]. In recent years, in connection with the rapid development of experimental technology, previously very problematic studies that are of key importance for the stereochemistry of heterocycles have become rather more possible and routine.…”

Cell Wall Integrity Pathway Involved in Morphogenesis, Virulence and Antifungal Susceptibility in Cryptococcus neoformans

“…In a recent work published by Sellers-Moya et al the authors showed how imidazoles, and particularly clotrimazole, trigger more changes in MAPK phosphorylation than triazoles, which implies an activation of MAPKs Hog1 and Slt2 high HOG and CWI pathways, respectively in S. cerevisiae [90]. They showed that clotrimazole induces a high level of ROS in this yeast compared with fluconazole.…”

“…In addition, resistance to this drug depends on both routes, HOG and CWI pathways. Regarding CWI they observed the activation by clotrimazole of Slt2 does not depend on pkc1 [90]. Amphotericin B alters the permeability of the fungal membrane by forming ion channels.…”