Regulation of the inositol transporter Itr1p by hydrogen peroxide in Saccharomyces cerevisiae
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ORIGINAL PAPER
Regulation of the inositol transporter Itr1p by hydrogen peroxide in Saccharomyces cerevisiae Tomás Santos1 · H. Susana Marinho1,2 · Luisa Cyrne1,2 Received: 12 April 2018 / Revised: 25 September 2018 / Accepted: 28 September 2018 © Springer-Verlag GmbH Germany, part of Springer Nature 2018
Abstract Myo-inositol is a precursor of several membrane phospholipids and sphingolipids and plays a key role in gene regulation in Saccharomyces cerevisiae (S. cerevisiae). Here, we tested whether H 2O2 was affecting the levels of the inositol transporters and thus inositol uptake. In S. cerevisiae cells adapted to H2O2 Itr1–GFPp accumulated in the plasma membrane until 20 min, concomitantly with an inhibition of its internalization. Exposure to H2O2 did not alter Itr2–GFPp cellular levels and induced only an 8% decrease at 10 min in the plasma membrane. Therefore, decreased inositol intracellular levels are not caused by decreased levels of inositol transporters in the plasma membrane. However, results show that H2O2 adaptation affects Itr1p turnover and, consequently, H2O2-adapted yeast cells display an inositol transporter phenotype comparable to cells grown in the absence of inositol in growth medium, i.e. accumulation in the plasma membrane and decreased degradation. Keywords Redox signaling · Hydrogen peroxide · Saccharomyces cerevisiae · Myo-inositol transporters · Inositol · Adaptation Abbreviations ANOVA One-way analysis of variance Art5p Arrestin-related trafficking adaptors GFP–Opi1p Fusion between green fluorescence protein and Opi1 protein H2O2 Hydrogen peroxide INO1 Inositol-1-phosphate synthase gene INO2 Ino2p gene ITR1 Myo-inositol transporter 1 gene Itr1–GFPp Fusion between Itr1 protein and green fluorescence protein ITR2 Myo-inositol transporter 2 gene
Communicated by Olaf Kniemeyer. * Luisa Cyrne [email protected] Tomás Santos [email protected] H. Susana Marinho [email protected] 1
Centro de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, 1749‑016 Lisbon, Portugal
Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, 1749‑016 Lisbon, Portugal
2
Itr2–GFPp Fusion between Itr2 protein and green fluorescence protein UASINO Inositol upstream activating sequence Rsp5p E3 ubiquitin ligase that conjugates ubiquitin to proteins to target them for degradation SC Synthetic complete medium
Introduction Myo-inositol (inositol) is essential for cellular structure and intracellular signalling regulation in all eukaryotes (Strahl and Thorner 2007; Berridge and Irvine 1989; Tsui and York 2010). In yeast, inositol is an essential precursor for the biosynthesis of phosphatidylinositol (PI) which can be incorporated in PI-derived molecules, such as complex sphingolipids and glycosylphosphatidylinositol anchors (Carman and Han 2011; Carman and Henry 1999; Henry et al. 2012; Cowart and Obeid 2007; Paulick and Bertozzi 2008). Inositol has also a major role in the transcriptional regulation of numerous genes
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