The MAPK Hog1p Modulates Fps1p-dependent Arsenite Uptake and Tolerance in Yeast

Abstract

Arsenic is widely distributed in nature and all organisms possess regulatory mechanisms to evade toxicity and acquire tolerance. Yet, little is known about arsenic sensing and signaling mechanisms or about their impact on tolerance and detoxification systems. Here, we describe a novel role of the S. cerevisiae mitogen-activated protein kinase Hog1p in protecting cells during exposure to arsenite and the related metalloid antimonite. Cells impaired in Hog1p function are metalloid hypersensitive, whereas cells with elevated Hog1p activity display improved tolerance. Hog1p is phosphorylated in response to arsenite and this phosphorylation requires Ssk1p and Pbs2p. Arsenite-activated Hog1p remains primarily cytoplasmic and does not mediate a major transcriptional response. Instead, hog1⌬ sensitivity is accompanied by elevated cellular arsenic levels and we demonstrate that increased arsenite influx is dependent on the aquaglyceroporin Fps1p. Fps1p is phosphorylated on threonine 231 in vivo and this phosphorylation critically affects Fps1p activity. Moreover, Hog1p is shown to affect Fps1p phosphorylation. Our data are the first to demonstrate Hog1p activation by metalloids and provides a mechanism by which this kinase contributes to tolerance acquisition. Understanding how arsenite/antimonite uptake and toxicity is modulated may prove of value for their use in medical therapy.

Novel stress responses facilitate Saccharomyces cerevisiae growth in the presence of the monocarboxylate preservatives

“…Resistances are induced to Acetic, but not propionic, sorbic or benzoic acids [19] Propionic, sorbic, benzoic acids, but not acetic acid [1,7] Key induced signalling pathway and activity HOG pathway signalling, leading to Hog1p MAP kinase activation [19] No evidence for a signalling pathway involvement; acid anion may act directly on the War1p transcription factor [13] Key action/target of the induced activity needed for the acquisition of resistance Phosphorylation of Fps1p, leading to Fps1p endocytosis [20] PDR12 gene induction, leading to an elevated plasma membrane level and activity of the Pdr12p ABC transporter [23,28] Probable related stress response Hog1p-directed downregulation of metalloid entry through the Fps1p channel [40] War1p activation in media containing leucine, methionine or phenylalanine as sole nitrogen source [9] Normal physiological function of the major target of the response Fps1p is the major glycerol channel of the plasma membrane, important for osmoadaptation [10,22,38] Pdr12p is also involved in export of the potentially toxic aromatic and branched-chain organic acids produced during amino acid catabolism [9] (2), a MAP kinase constitutively bound to, and that directly phosphorylates, Fps1p (3). This phosphorylation provides the signal for Fps1p endocytosis and degradation, eliminating this source of the acetic acid entry to the cell.…”

Arsenic Toxicity to Saccharomyces cerevisiae Is a Consequence of Inhibition of the TORC1 Kinase Combined with a Chronic Stress Response

“…A number of mechanisms exist for detoxification, probably because arsenic has always been widespread in the environment. These involve reduction of influx through the aquaglyceroporin Fps1p Thorsen et al, 2006); sequestration into the vacuole in the form of glutathione conjugates, metallothionein, and other metal/protein complexes; and active extrusion (Ghosh et al, 1999). In yeast, genome-wide analysis of the transcription patterns in response to arsenic revealed a complex network of transcription factors controlling the expression of several hundred genes (Haugen et al, 2004;Wysocki et al, 2004;Thorsen et al, 2007).…”

“…In yeast, genome-wide analysis of the transcription patterns in response to arsenic revealed a complex network of transcription factors controlling the expression of several hundred genes (Haugen et al, 2004;Wysocki et al, 2004;Thorsen et al, 2007). Mitogen-activated protein kinases mediate protective responses involving AP-1-and AP-1-like transcription factors in higher eukaryotes and in fungi (Cavigelli et al, 1996;Rodriguez-Gabriel and Russell, 2005;Thorsen et al, 2006).…”

Presence of the Fps1p aquaglyceroporin channel is essential for Hog1p activation, but suppresses Slt2(Mpk1)p activation, with acetic acid stress of yeast

“…Activated in response to hyperosmotic stress (Hedfalk et al, 2004;Pettersson et al, 2005;Tamas et al, 2003), arsenite (Thorsen et al, 2006) and acetic acid (Mollapour & Piper, 2006), it induces physiologically very different responses under these diverse conditions of stress. Only with acetic acid, not hyperosmotic or arsenite stress, has the active Hog1p been shown to trigger endocytosis of Fps1p, the major plasma membrane channel for glycerol, arsenite and undissociated acetic acid (Mollapour & Piper, 2007).…”

“…This is analogous to the way that Hog1p alleviates salt stress, a situation where this same MAP kinase -but now the Hog1p subfraction in association with certain plasma membrane ion transporters (Nha1p and Tok1p) -generates an instant relief of NaCl stress [Hog1p stimulation of Nha1p Na + /H + antiporter activity following salt stress is practically instant, preceding any Hog1p-dependent changes in gene transcription (Proft & Struhl, 2004)]. Hog1p also directly phosphorylates Fps1p in the response to arsenite stress, though unlike with acetic acid treatment, it appears that here it causes the closure, not the destabilization, of this channel (Thorsen et al, 2006). Fps1p channel closure is also triggered by osmostress, but this is a turgor-, rather than a Hog1p-mediated event and extremely rapid [Fps1p-mediated glycerol flux diminishes within seconds following a shift to high osmolarity and increases over an apparently similar timescale when cells are then shifted to low osmolarity (Luyten et al, 1995;Tamas et al, 1999)].…”