THE ETS FAMILY MEMBER ERM CONTAINS AN ALPHA-HELICAL ACIDIC ACTIVATIONDOMAIN THAT CONTACTS TAFII60

Transcription factors are modular entities built up of: discrete domai ns, some devoted to DNA binding and others permitting transcriptional modulation. The structure of DNA binding domains has been thoroughly i nvestigated and structural classes clearly defined, In sharp contrast, the structural constraints put on transactivating regions, if any, ar e mostly unknown, Our investigations focus on ERM, a eukaryotic transc ription factor of the ETS family, We have previously shown that ERM ha rbours two transactivating domains (TADs) with distinct functional fea tures: AD1 lies in the first 72 amino acids of ERM, while AD2 sits in the last 62, Here we show that AD1 is a bona fide acidic TAD, for it a ctivated transcription in yeast cells, while AD2 did not, AD1 contains a 20 amino acid stretch predicted to form an alpha-helix that is foun d unchanged in the related PEA3 and ER81 transcription factors, Circul ar dichroism analysis revealed that a 32 amino acid peptide encompassi ng this region is unstructured in water but folds into a helix when th e hydrophobic solvent trifluoroethanol is added, The isolated helix wa s sufficient to activate transcription and mutations predicted to disr upt it dramatically affected AD1-driven transactivation, whereas mutat ions decreasing its acidity had more gentle effects, A phenylalanine r esidue within the helix was particularly sensitive to mutations, Final ly we observed that ERM bound TAFII60 via AD1 and bound TBP and TAFII4 0, presumably via other activation domains.