Interaction of the transcription factor Sp1 with the nuclear pore protein p62 requires the C-terminal domain of p62.
The transcription factor Sp1 plays an important role in the expression of many cellular genes. In studies of proteins that associate with Sp1, a 62-kDa glycoprotein was found in immunoprecipitates of Sp1. This protein was detected in these immunoprecipitates by the monoclonal antibody, RL2, which was originally raised against nuclear pore proteins but was subsequently found to recognize an epitope that contains O-linked N-acetylglucosamine (O-GlcNAc). The association of this protein with Sp1 could be blocked by SDS denaturation of the protein complex. Western blot analysis of the Sp1 immunoprecipitate using antibodies to p62 nucleoporin indicated that this nuclear pore protein associates with Sp1. Furthermore, immunoprecipitation of p62 nucleoporin resulted in the coprecipitation of Sp1. Recombinant p62, expressed as a GST-fusion protein using a vaccinia virus system, also interacted with both recombinant and native Sp1. This interaction between p62 and Sp1 required the C-terminus of p62 and the C-terminus was able to bind Sp1, albeit less efficiently than native p62. A mammalian two-hybrid interaction assay was devised in which p62 was fused to the Gal4 DNA-binding domain. This system also indicated that p62, through its C-terminus, interacts with Sp1 in the living cell. We propose that this interaction of a nuclear pore protein with Sp1 may reflect the nuclear organization required to bring transcribable DNA in contact with the transcription factors.
Han
I
I
Department of Medicine, University of Alabama at Birmingham 35294, USA.
Roos
M D
MD
Kudlow
J E
JE
D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002478:Cells, Cultured; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins; D010446:Peptide Fragments / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D013501:Surface-Active Agents / Q000494:pharmacology
D008562:Membrane Glycoproteins; D028861:Nuclear Pore Complex Proteins; D010446:Peptide Fragments; D016329:Sp1 Transcription Factor; D013501:Surface-Active Agents; C049312:nuclear pore protein p62; D012967:Sodium Dodecyl Sulfate