Gene activation by recruitment of the RNA polymerase II holoenzyme.
The single amino acid "P" (potentiator) mutation in the holoenzyme component GAL11 creates an interaction between that protein and the dimerization region of GAL4. That interaction triggers strong gene activation when the GAL4 fragment is tethered to DNA. Here we show that, among a series of variants of the GAL4 dimerization region and different GAL11P alleles, the strength of the interaction as quantitated in vitro correlates with the degree of activation in vivo; swapping the protein fragments bearing the GAL4 dimerization region and the GAL11P mutation such that the latter is tethered to DNA and the former is attached to the holoenzyme does not diminish gene activation; gene activation in this system is squelched by overproduction of either a fragment bearing the GAL4 dimerization region or a fragment of GAL11 bearing a P mutation; and neither GAL11 nor GAL11P is a target of an acidic activating region. These results argue that the GAL4-GAL11P interaction triggers gene activation simply by recruiting the holoenzyme to DNA. Consistent with this view, we also show that fusion of LexA to another holoenzyme component, SRB2, creates an activator, and that an SRB2 mutant predicted on genetic grounds to interact especially efficiently with a holoenzyme containing a specific mutant form of polymerase also activates more efficiently when tethered to DNA.
Farrell
S
S
Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA.
Simkovich
N
N
Wu
Y
Y
Barberis
A
A
Ptashne
M
M
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism*; D005786:Gene Expression Regulation; D056892:Mediator Complex; D008957:Models, Genetic; D009154:Mutation; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation; D000099160:LexA Repressor Protein
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D056892:Mediator Complex; D010446:Peptide Fragments; D012319:RNA Polymerase II; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D012697:Serine Endopeptidases; D015534:Trans-Activators; D014157:Transcription Factors; D000099160:LexA Repressor Protein; C058337:GAL11 protein, S cerevisiae; C052130:GAL4 protein, S cerevisiae; C074748:SRB2 protein, S cerevisiae