Molecular basis of the interactions between the p73 N terminus and p300: effects on transactivation and modulation by phosphorylation.
The transcription factor p73 belongs to the p53 family of proteins and can transactivate a number of target genes in common with p53. Here, we characterized the interaction of the p73 N terminus with four domains of the transcriptional coactivator p300 and with the negative regulator Mdm2 by using biophysical and cellular measurements. We found that, like p53, the N terminus of p73 contained two distinct transactivation subdomains, comprising residues 10-30 and residues 46-67. The p73 N terminus bound weakly to the Taz1, Kix, and IBiD domains of p300 but with submicromolar affinity for Taz2, in contrast to previous reports. We found weaker binding of the p73 N terminus to the p300 domains in vitro correlated with a significant decrease in transactivation activity in a cell line for the QS and T14A mutants, and tighter binding of the phosphomimetic T14D in vitro correlated with an increase in vivo. Further, we found that phosphorylation of T14 increased the affinity of the p73 N terminus for Taz2 10-fold. The phosphomimetic p73alpha T14D caused increased levels of transactivation.
Burge
Sarah
S
Medical Research Council Centre for Protein Engineering, Hills Road, Cambridge CB2 0QH, United Kingdom.
Teufel
Daniel P
DP
Townsley
Fiona M
FM
Freund
Stefan M V
SM
Bycroft
Mark
M
Fersht
Alan R
AR
D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D050881:E1A-Associated p300 Protein / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D016415:Sequence Alignment; D013816:Thermodynamics; D015533:Transcriptional Activation / Q000235:genetics*; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000737:chemistry / Q000378:metabolism
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C000606460:TP73 protein, human; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D051028:bcl-2-Associated X Protein; D050881:E1A-Associated p300 Protein