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Journal of Bacteriology, May 2001, p. 3098-3107, Vol. 183, No. 10
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.10.3098-3107.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Tagging Morphogenetic Genes by Insertional Mutagenesis in the Yeast Yarrowia lipolytica

Mathias Richard,1,* Raymundo Rosas Quijano,1,dagger Samira Bezzate,1 Florence Bordon-Pallier,2 and Claude Gaillardin1

Laboratoire de Génétique Moléculaire et Cellulaire, Institut National Agronomique Paris-Grignon, UMR-INRA216, URA-CNRS1925, 78850 Thiverval-Grignon,1 and Hoechst-Marion-Roussel, Aventis Pharma, 93235 Romainville Cedex,2 France

Received 1 December 2000/Accepted 26 February 2001

The yeast Yarrowia lipolytica is distantly related to Saccharomyces cerevisiae, can be genetically modified, and can grow in both haploid and diploid states in either yeast, pseudomycelial, or mycelial forms, depending on environmental conditions. Previous results have indicated that the STE and RIM pathways, which mediate cellular switching in other dimorphic yeasts, are not required for Y. lipolytica morphogenesis. To identify the pathways involved in morphogenesis, we mutagenized a wild-type strain of Y. lipolytica with a Tn3 derivative. We isolated eight tagged mutants, entirely defective in hyphal formation, from a total of 40,000 mutants and identified seven genes homologous to S. cerevisiae CDC25, RAS2, BUD6, KEX2, GPI7, SNF5, and PPH21. We analyzed their abilities to invade agar and to form pseudomycelium or hyphae under inducing conditions and their sensitivity to temperature and to Calcofluor white. Chitin staining was used to detect defects in their cell walls. Our results indicate that a functional Ras-cyclic AMP pathway is required for the formation of hyphae in Y. lipolytica and that perturbations in the processing of extracellular, possibly parietal, proteins result in morphogenetic defects.

* Corresponding author. Mailing address: Laboratoire de Génétique Moléculaire et Cellulaire, BP01, 78850 Thiverval-Grignon, France. Phone: 33 1 30 81 54 43. Fax: 33 1 30 81 54 57. E-mail: richard{at}grignon.inra.fr.

dagger Present address: Centro de Investigacion y de Estudios Avanzados, Instituto Politecnico Nacional, Irapuato Unit, Irapuato 36500, Guanajuato, Mexico.

Journal of Bacteriology, May 2001, p. 3098-3107, Vol. 183, No. 10
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.10.3098-3107.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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