Overexpression of YAP2, coding for a new yAP protein, and YAP1 in Saccharomyces cerevisiae alleviates growth inhibition caused by 1,10-phenanthroline.

P Bossier,L Fernandes,D Rocha,C Rodrigues-Pousada

doi:10.1016/s0021-9258(19)49510-0

P Bossier, L Fernandes + Show 2 more

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https://doi.org/10.1016/s0021-9258(19)49510-0

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Abstract

A new member of the yeast yAP family, designated YAP2, has been isolated and characterized. The protein displays a high homology with the DNA-binding domain of yAP-1, which contains a basic DNA-binding domain and leucine zipper motif and binds in vitro to the same cis-element. Growth arrest of yeast caused by low concentrations of 1,10-phenanthroline, resulting in zinc and/or iron deprivation, is overcome by over-expression of YAP1 or YAP2. In fact, yeast cells over-expressing YAP1 or YAP2 display pleiotropic drug resistance. On the other hand, a yap2 null mutant has an increased thermotolerance under starvation conditions caused by 1,10-phenanthroline. The latter mutant might become an excellent tool in the study of pathways leading toward thermotolerance acquisition.

Highlights

Recipient of a postdoctoral fellowshipof the Calouste GulbenkianFoundation. $ Supported by the EEC Erasmus program. ll To whom correspondence should be addressed
From the Znstituto Gulbenkian de Ciincia, Laboratorio Genitica Molecular, Rua da Quinta Grande, Apartado 14, 2781 Oeiras Codex, Portugal
The pro- clone yeast genes that at high copy number would be able to tein displays a high homology with the DNA-binding overcome growth arrest caused by 1,lO-phenanthroline

Summary

Recipient of a postdoctoral fellowshipof the Calouste Gulbenkian

Foundation. $ Supported by the EEC Erasmus program. ll To whom correspondence should be addressed. Localized at position -157 relative to the main ATG codon (Fig. 3) This implies that a small ORF found upstreamof the Cloning of the YAP1 and YAP2 Gene-In solidified minimal mainATG(uORF)issituatedwithinthe5”untranslated medium containing 1,lO-phenanthrolin(e10 pglml), the yeast leader sequence of this new gene. Yeast cells regain the DNA-binding domain,we designated thegene in thesubclone possibility to grow inthepresence of 1,lO-phenanthroline pPR62.lG YAP2 and its gene product YAP-2 This designawhen sufficient zincor iron is added to the medium to sattuioranties supported by the fact that whole cell extracts from the chelating agent (results not showAn).yeast library in the YAPl andYAP2 overexpressing strains overproduce aprotein vector pMA3a (Crouzet and Tuite, 1987) was used to trans- that binds ivnitro to the TGACT-containing ARE (Harshman form the strain W303a (see “Materials and Methods”).

G M GATTCACTCCCGICCACA

HOMOLOGY IN bZIP MOTIF

HOMOLOGY IN C R 2

DISCUSSION