Different Smad2 partners bind a common hydrophobic pocket in Smad2 via a defined proline-rich motif.
Transforming growth factor-beta (TGF-beta)/activin-induced Smad2/Smad4 complexes are recruited to different promoter elements by transcription factors, such as Fast-1 or the Mix family proteins Mixer and Milk, through a direct interaction between Smad2 and a common Smad interaction motif (SIM) in the transcription factors. Here we identify residues in the SIM critical for Mixer-Smad2 interaction and confirm their functional importance by demonstrating that only Xenopus and zebrafish Mix family members containing a SIM with all the correct critical residues can bind Smad2 and mediate TGF-beta-induced transcriptional activation in vivo. We identify significant sequence similarity between the SIM and the Smad-binding domain (SBD) of the membrane-associated protein SARA (Smad anchor for receptor activation). Molecular modelling, supported by mutational analyses of Smad2 and the SIM and the demonstration that the SARA SBD competes directly with the SIM for binding to Smad2, indicates that the SIM binds Smad2 in the same hydrophobic pocket as does the proline-rich rigid coil region of the SARA SBD. Thus, different Smad2 partners, whether cytoplasmic or nuclear, interact with the same binding pocket in Smad2 through a common proline-rich motif.
Randall
Rebecca A
RA
Laboratory of Developmental Signalling and Biomolecular Modelling Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, UK.
Germain
Stéphane
S
Inman
Gareth J
GJ
Bates
Paul A
PA
Hill
Caroline S
CS
D016475:3T3 Cells; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011392:Proline / Q000737:chemistry; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D051899:Smad2 Protein; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014981:Xenopus / Q000235:genetics / Q000378:metabolism; D029867:Xenopus Proteins; D015027:Zebrafish / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D051899:Smad2 Protein; C497785:Smad2 protein, Xenopus; C497783:Smad2 protein, mouse; D015534:Trans-Activators; D014157:Transcription Factors; D029867:Xenopus Proteins; C492108:ZFYVE9 protein, Xenopus; D029961:Zebrafish Proteins; C405705:mixer protein, Xenopus; C499590:mixl1 protein, zebrafish; C497786:smad2 protein, zebrafish; D004247:DNA; D011392:Proline