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Applied and Environmental Microbiology, May 2002, p. 2155-2160, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2155-2160.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Impaired Cutinase Secretion in Saccharomyces cerevisiae Induces Irregular Endoplasmic Reticulum (ER) Membrane Proliferation, Oxidative Stress, and ER-Associated Degradation

C. M. J. Sagt,¹ W. H. Müller,² L. van der Heide,² J. Boonstra,^2* A. J. Verkleij,² and C. T. Verrips^2,3

DSM-Gist, 2600 MA Delft,¹ Department of Molecular Cell Biology and Institute of Biomembranes, Utrecht University, 3584 CH Utrecht,² Unilever Research Laboratory Vlaardingen, 3133 AT Vlaardingen, The Netherlands³

Received 9 November 2001/ Accepted 28 January 2002

Impaired secretion of the hydrophobic CY028 cutinase invokes an unfolded protein response (UPR) in Saccharomyces cerevisiae cells. Here we show that the UPR in CY028-expressing S. cerevisiae cells is manifested as an aberrant morphology of the endoplasmic reticulum (ER) and as extensive membrane proliferation compared to the ER morphology and membrane proliferation of wild-type CY000-producing S. cerevisiae cells. In addition, we observed oxidative stress, which resulted in a 21-fold increase in carbonylated proteins in the CY028-producing S. cerevisiae cells. Moreover, CY028-producing S. cerevisiae cells use proteasomal degradation to reduce the amount of accumulated CY028 cutinase, thereby attenuating the stress invoked by CY028 cutinase expression. This proteasomal degradation occurs within minutes and is characteristic of ER-associated degradation (ERAD). Our results clearly show that impaired secretion of the heterologous, hydrophobic CY028 cutinase in S. cerevisiae cells leads to protein aggregation in the ER, aberrant ER morphology and proliferation, and oxidative stress, as well as a UPR and ERAD.

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* Corresponding author. Mailing address: Faculty of Biology, Department of Molecular Cell Biology, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands. Phone: 31 30 2533189. Fax: 31 30 2513655. E-mail: J.Boonstra{at}bio.uu.nl.

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Applied and Environmental Microbiology, May 2002, p. 2155-2160, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2155-2160.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

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