tailieunhanh - Báo cáo khoa học: The Saccharomyces cerevisiae vacuolar acid trehalase is targeted at the cell surface for its physiological function

Previous studies in the yeastSaccharomyces cerevisiaehave proposed a vac-uolar localization for Ath1, which is difficult to reconcile with its ability to hydrolyze exogenous trehalose. We used fluorescent microscopy to show that the red fluorescent protein mCherry fused to the C-terminus of Ath1, although mostly localized in the vacuole, was also targeted to the cell sur-face. | The Saccharomyces cerevisiae vacuolar acid trehalase is targeted at the cell surface for its physiological function Susu He1 2 3 Kerstin Bystricky4 5 Sebastien Leon6 Jean M. Francois1 2 3 and Jean L. Parrou1 2 3 1 University of Toulouse INSA UPS INP INRA France 2 INRA-UMR 792 Ingenierie des Systemes Biologiques et precedes Toulouse France 3 CNRS-UMR 5504 Toulouse France 4 Laboratoire de Biologie Moleculaire Eucaryote University of Toulouse France 5 CNRS-UMR5099 Toulouse France 6 Institut Jacques Monod UMR7592 CNRS Universite Paris Diderot France Keywords acid trehalase cell surface fluorescence microscopy Saccharomyces cerevisiae secretion Correspondence J. M. Francois University of Toulouse INSA UPS INP INRA 135 Avenue de Rangeuil F-31077 Toulouse France Fax 33 5 6155 9400 Tel 33 5 6155 9492 E-mail franjm@ Received 6 May 2009 revised 9 June 2009 accepted 21 July 2009 doi Previous studies in the yeast Saccharomyces cerevisiae have proposed a vacuolar localization for Ath1 which is difficult to reconcile with its ability to hydrolyze exogenous trehalose. We used fluorescent microscopy to show that the red fluorescent protein mCherry fused to the C-terminus of Ath1 although mostly localized in the vacuole was also targeted to the cell surface. Also hybrid Ath1 truncates fused at their C-terminus with the yeast internal invertase revealed that a 131 amino acid N-terminal fragment of Ath1was sufficient to target the fusion protein to the cell surface enabling growth of the suc2D mutant on sucrose. The unique transmembrane domain appeared to be indispensable for the production of a functional Ath1 and its removal abrogated invertase secretion and growth on sucrose. Finally the physiological significance of the cell-surface localization of Ath1 was established by showing that fusion of the signal peptide of invertase to N-terminal truncated Ath1 allowed the athlD mutant to grow on trehalose whereas the signal sequence of .