Glycogen synthase kinase-3beta binds to E2F1 and regulates its transcriptional activity.
GSK3beta and E2F1 play an important role in the control of proliferation and apoptosis. Previous work has demonstrated that GSK3beta indirectly regulates E2F activity through modulation of cyclin D1 levels. In this work we show that GSK3beta phosphorylates human E2F1 in vitro at serine 403 and threonine 433, both residues localized at its transactivation domain. This phosphorylation was not detected in vivo. However, co-immunoprecipitation experiments do reveal in vivo binding of these proteins. Moreover, uninhibitable and catalitycally inactive GSK3beta forms inhibit the transcriptional activity of a fusion protein containing E2F1 transactivation domain. Both forms of GSK3beta inhibit E2F1 with similar efficiency. Interestingly the effect was independent of the mutation of serine 403 and threonine 433 to alanine. This suggests that this transcriptional modulation is independent of GSK3beta kinase activity and phosphorylation state of serine 403 and threonine 433. The re-targeting of these GSK3beta forms to the nucleus results in a higher capacity to regulate E2F1 transcriptional activity. Depletion of the levels of GSK3beta protein using siRNA activates E2F1 transcriptional activity. The data presented in this study offer a new mechanism of regulation of E2F1 by direct binding of GSK3beta to its transactivation domain.
García-Alvarez
Gisela
G
Departament de Bioquímica i Biologia Molecular, Facultat de Farmàcia, Universitat de Barcelona. Av. Diagonal 643, E-08028 Barcelona, Catalunya, Spain.
Ventura
Verònica
V
Ros
Oriol
O
Aligué
Rosa
R
Gil
Joan
J
Tauler
Albert
A
D002460:Cell Line; D050687:E2F1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D010768:Phosphoserine / Q000378:metabolism; D010769:Phosphothreonine / Q000378:metabolism; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D034741:RNA, Small Interfering; D015534:Trans-Activators; D010769:Phosphothreonine; D010768:Phosphoserine; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D038362:Glycogen Synthase Kinase 3