p90 Ribosomal S6 Kinase 1 (RSK1) and the Catalytic Subunit of Protein Kinase A (PKA) Compete for Binding the Pseudosubstrate Region of PKAR1α 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, PKARI alpha, 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 PKARI alpha. Substitution of the activation loop phosphorylation site (Ser-221) in the NTK with the negatively charged Asp residue abrogated the association between RSK1 and PKARI alpha. This explains the lack of an interaction between active RSK1 and PKARI alpha. Full-length RSK1 bound to PKARI alpha 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 PKARI alpha. Overexpressed RSK1 dissociated PKAc from PKARI alpha, increasing PKAc activity, whereas silencing of RSK1 increased PKAc/PKARI alpha interactions and decreased PKAc activity. Unlike PKAc, which requires Arg-95 and -96 in the pseudosubstrate region of PKARI alpha for their interactions, RSK1/PKARI alpha association requires all four Arg residues (Arg-93-96) in the pseudosubstrate site of PKARI alpha. A peptide (Wt-PS) corresponding to residues 91-99 of PKARI alpha competed for binding of RSK1 with PKARI alpha both in vitro and in intact cells. Furthermore, peptide Wt-PS (but not control peptide Mut-PS), by dissociating RSK1 from PKARI alpha, activated RSK1 in the absence of any growth factors and protected cells from apoptosis. Thus, by competing for binding to the pseudosubstrate region of PKARI alpha, RSK1 regulates PKAc activity in a cAMP-independent manner, and PKARI alpha by associating with RSK1 regulates its activation and its biological functions.