STUDY OF EFFECTS FROM DELETION OF AGT1 AND ATH1 UPON THERMAL STRESS IN SACCHAROMYCES CEREVISIAE
R.S.S,Magalhães¹ ,J. F., de Mesquita²,E.C.A. Eleutherio ¹,
¹ Programa de Pós-graduação de Bioquimica, Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro; ² Departamento de Genética e Biologia Molecular, Instituto Biomédico, Universidade Federal do Estado do Rio de Janeiro.
Saccharomyces cerevisiae accumulates trehalose in response to stress. This sugar protects biomolecules, increasing cell tolerance. To stabilize membranes, trehalose must be present on both sides of the bilayer. After a stress, trehalose is degraded. The goal of this work was to elucidate the molecular mechanism by which trehalose, synthesized in the cytosol, is transported to the outside of bilayer and degraded during stress recovery. Thus, we used mutants deficient in trehalose transport (Dagt1), cytosolic trehalase (Dnth1) or in periplasmic/vacuolar trehalase (Dath1). Cells growing exponentially on glucose at 28°C were directly exposed to 51°C/ 8 min or previously submitted to an adaptive heat-treatment at 40 °C/60 min and then exposed to heat stress. In some cases, immediately after 40 °C, cells were back to 28°C/1 h before being stressed. For each condition, viability (plating on solid glucose-medium) and trehalose accumulation (alcohol extraction and quantification by HPLC) were analyzed. The treatment at 40°C, which induces trehalose synthesis, increased the tolerance of all strains. Dnth1 showed the highest tolerance to heat stress. This mutant accumulated the highest concentration of trehalose over the others strain. Both Dnth1 and Dath1 presented a higher tolerance to heat stress than wild-type when they were shifted back to 28°C, which seems to be related to the high trehalose levels that remained even after the stress recovery. Dagt1 and wild-type strains showed the same profile of accumulation of trehalose, although the tolerance of the mutant was lower. This result may be related to the absence of trehalose in the outside of the lipid bilayer. These results suggest that Agt1 exports trehalose synthesized in the cytosol to protect the outside of the plasma membrane. During stress recovery, those external trehalose molecules seem to be degraded by Ath1.
Financial Support: CAPES, CNPq Area: Biochemistry
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