Abstract The role of trehalose as cell protector against oxidative stress induced by H 2 O 2 has been studied in Sac- charomyces cerevisiae mutants in which the two trehalase genes ATH1 and NTH1 are deleted. The addition of low H 2 O 2 concentrations to proliferating cultures of either strain did not harm cell viability and induced a marked ac- tivity to Nth1p, but with no significant level of trehalose accumulation. This pattern was reversed after a more se- vere H 2 O 2 treatment that caused drastic cell killing. The most severe phenotype corresponded to the ∆nth1 mutant. Under these conditions, the increase in Nth1p was abol- ished and a three-fold rise in trehalose content was recorded concomitant with activation of the trehalose syn- thase complex. The behavior of the double-disruptant ∆ath1∆nth1 mutant was identical to that of wild-type cells, although in exponential cultures Ath1p activity was virtually undetectable upon exposure to H 2 O 2 . Further- more, these strains displayed an adaptive response to ox- idative stress that was independent of intracellular tre- halose synthesis. Our data strongly suggest that trehalose storage in budding yeasts is not an essential protectant in cell defense against oxidative challenge. Keywords H 2 O 2 · Oxidative stress · Trehalose · Trehalase · Yeasts Introduction In the yeast Saccharomyces cerevisiae, the stress-respon- sive machinery is strictly dependent on the expression of a family of heat-shock proteins, whose counterpart genes contain in their promoter either the heat-shock element (HSE) or the stress-responsive element (STRE, CCCT) (Lindquist and Craig 1988; Estruch 2000). In turn, a sub- stantial accumulation of intracellular trehalose under stress challenges has been convincingly demonstrated to act as an important contributory factor to resistance (Wiemken 1990; Thevelein 1996; Argüelles 2000). Tre- halose is a non-reducing glucose disaccharide that seems to play a dual role: (1) preserving the integrity of the plasma membrane by substituting water and binding to the polar head groups of phospholipids (Crowe et al. 1984), and (2) functioning as a “chaperone” by stabilizing proteins in their native state during injury as well as by suppressing the aggregation of denatured proteins (Singer and Lindquist 1998). In addition, trehalose hydrolysis provides the energy necessary for correct renaturation of proteins during stress recovery (Singer and Lindquist 1998; Nwaka and Holzer 1998). Trehalose degradation in S. cerevisiae is exclusively confined to the enzyme trehalase (E.C. 3.2.1.28), of which two types are present. The first is a vacuolar enzyme (Ath1p), subject to glucose repression, whose optimum pH for activity is acidic (4.0–5.0) (San Miguel and Argüelles 1994; Destruelle et al. 1995; Thevelein 1996). This trehalase lacks any evident physiological role, al- though Ath1p may be responsible for the utilization and uptake of exogenous trehalose (Nwaka et al. 1996). The other trehalase, the neutral cytosolic enzyme (Nth1p), is well characterized. NTH1p has been cloned and mapped on chromosome IV; the enzyme exhibits maximal activity at pH~7.0, and is regulated by a cAMP-dependent phos- phorylation process (Londesborough and Varimo 1984; Kopp et al. 1993; Nwaka and Holzer 1998). A third puta- tive trehalase gene (NTH2) has been discovered (Nwaka et al. 1995), but it lacks both neutral and acid trehalase ac- tivity and has no detetectable influence on the intracellu- lar content of trehalose in intact cells (Nwaka et al. 1995). Nth1p carries out trehalose mobilization in different physiological situations and contains three STRE (CCCT) elements in its promoter region (Nwaka et al. 1995). Ac- Yolanda Pedreño · Jose V. Gimeno-Alcañiz · Emilia Matallana · Juan-Carlos Argüelles Response to oxidative stress caused by H 2 O 2 in Saccharomyces cerevisiae mutants deficient in trehalase genes Received: 14 October 2001 / Revised: 5 February 2002 / Accepted: 27 February 2002 / Published online: 4 April 2002 ORIGINAL PAPER Y. Pedreño · J.-C. Argüelles (✉) Area de Microbiología, Facultad de Biología. Universidad de Murcia. 30071 Murcia, Spain e-mail: arguelle@um.es, Tel.: +34-968-367131, Fax: +34-968-363963, J.V. Gimeno-Alcañiz · E. Matallana Departamento de Bioquímica y Biología Molecular, Universitat de Valencia e Instituto de Agroquímica y Tecnología de Alimentos, CSIC. 46100 Burjassot, Valencia, Spain Arch Microbiol (2002) 177 : 494–499 DOI 10.1007/s00203-002-0418-2 © Springer-Verlag 2002