Expression of phospholipid biosynthetic genes in yeast requires activator protein INO2 which can bind to the UAS element inositol/choline-responsive element (ICRE) and trigger activation of target genes, using two separate transcriptional activation domains, TAD1 and TAD2. However, it is still unknown which cofactors mediate activation by TADs of INO2. Here, we show that multiple subunits of basal transcription factor TFIID (TBP-associated factors TAF1, TAF4, TAF6, TAF10 and TAF12) are able to interact in vitro with activation domains of INO2. Interaction was no longer observed with activation-defective variants of TAD1. We were able to identify two nonoverlapping regions in the N-terminus of TAF1 (aa 1-100 and aa 182-250) each of which could interact with TAD1 of INO2 as well as with TAD4 of activator ADR1. Specific missense mutations within TAF1 domain aa 182-250 affecting basic and hydrophobic residues prevented interaction with wild-type TAD1 and caused reduced expression of INO1. Using chromatin immunoprecipitation we demonstrated INO2-dependent recruitment of TAF1 and TAF6 to ICRE-containing promoters INO1 and CHO2. Transcriptional derepression of INO1 was no longer possible with temperature-sensitive TAF1 and TAF6 mutants cultivated under nonpermissive conditions. This result supports the hypothesis of Taf-dependent expression of structural genes activated by INO2.

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Journal Article

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Hintze S, Engelhardt M, van Diepen L, Witt E, Schüller HJ

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