Plasminogen activator inhibitor type 1 interacts with alpha3 subunit of proteasome and modulates its activity.
Plasminogen activator inhibitor type-1 (PAI-1), a multifunctional protein, is an important physiological regulator of fibrinolysis, extracellular matrix homeostasis, and cell motility. Recent observations show that PAI-1 may also be implicated in maintaining integrity of cells, especially with respect to cellular proliferation or apoptosis. In the present study we provide evidence that PAI-1 interacts with proteasome and affects its activity. First, by using the yeast two-hybrid system, we found that the α3 subunit of proteasome directly interacts with PAI-1. Then, to ensure that the PAI-1-proteasome complex is formed in vivo, both proteins were coimmunoprecipitated from endothelial cells and identified with specific antibodies. The specificity of this interaction was evidenced after transfection of HeLa cells with pCMV-PAI-1 and coimmunoprecipitation of both proteins with anti-PAI-1 antibodies. Subsequently, cellular distribution of the PAI-1-proteasome complexes was established by immunogold staining and electron microscopy analyses. Both proteins appeared in a diffuse cytosolic pattern but also could be found in a dense perinuclear and nuclear location. Furthermore, PAI-1 induced formation of aggresomes freely located in endothelial cytoplasm. Increased PAI-1 expression abrogated degradation of degron analyzed after cotransfection of HeLa cells with pCMV-PAI-1 and pd2EGFP-N1 and prevented degradation of p53 as well as IκBα, as evidenced both by confocal microscopy and Western immunoblotting.
Institute of Medical Biology, Polish Academy of Sciences, Lodz 93-232, Poland.
Joanna
J
Boncela
Patrycja
P
Przygodzka
Izabela
I
Papiewska-Pajak
Elzbieta
E
Wyroba
Magdalena
M
Osinska
Czeslaw S
CS
Cierniewski
D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins / Q000235:genetics / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D020672:I-kappa B Proteins; C000606204:NFKBIA protein, human; D017395:Plasminogen Activator Inhibitor 1; C051321:SERPINE1 protein, human; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D000072000:NF-KappaB Inhibitor alpha; C488165:PSMA3 protein, human; D046988:Proteasome Endopeptidase Complex