p90 ribosomal S6 kinase 1 (RSK1) and the catalytic subunit of protein kinase A (PKA) compete for binding the pseudosubstrate region of PKAR1alpha: role in the regulation of PKA and RSK1 activities.
Previously we showed that the inactive form of p90 ribosomal S6 kinase 1 (RSK1) interacts with the regulatory subunit, PKARIalpha, of protein kinase A (PKA), whereas the active RSK1 interacts with the catalytic subunit (PKAc) of PKA. Herein, we demonstrate that the N-terminal kinase domain (NTK) of RSK1 is necessary for interactions with PKARIalpha. Substitution of the activation loop phosphorylation site (Ser-221) in the NTK with the negatively charged Asp residue abrogated the association between RSK1 and PKARIalpha. This explains the lack of an interaction between active RSK1 and PKARIalpha. Full-length RSK1 bound to PKARIalpha with an affinity of 0.8 nm. The NTK domain of RSK1 competed with PKAc for binding to the pseudosubstrate region (amino acids 93-99) of PKARIalpha. Overexpressed RSK1 dissociated PKAc from PKARIalpha, increasing PKAc activity, whereas silencing of RSK1 increased PKAc/PKARIalpha interactions and decreased PKAc activity. Unlike PKAc, which requires Arg-95 and -96 in the pseudosubstrate region of PKARIalpha for their interactions, RSK1/PKARIalpha association requires all four Arg residues (Arg-93-96) in the pseudosubstrate site of PKARIalpha. A peptide (Wt-PS) corresponding to residues 91-99 of PKARIalpha competed for binding of RSK1 with PKARIalpha both in vitro and in intact cells. Furthermore, peptide Wt-PS (but not control peptide Mut-PS), by dissociating RSK1 from PKARIalpha, activated RSK1 in the absence of any growth factors and protected cells from apoptosis. Thus, by competing for binding to the pseudosubstrate region of PKARIalpha, RSK1 regulates PKAc activity in a cAMP-independent manner, and PKARIalpha by associating with RSK1 regulates its activation and its biological functions.
Gao
Xianlong
X
Department of Molecular Pharmacology and Therapeutics, Loyola University Chicago, Stritch School of Medicine, Maywood, Illinois 60153, USA.
Chaturvedi
Deepti
D
Patel
Tarun B
TB
D000818:Animals; D001665:Binding Sites; D020134:Catalytic Domain; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit / Q000235:genetics / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D051379:Mice; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000235:genetics / Q000378:metabolism*
D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; D007527:Isoenzymes; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D000242:Cyclic AMP; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D017868:Cyclic AMP-Dependent Protein Kinases