Autophagy is an evolutionarily conserved lysosomal pathway for degrading cytoplasmic proteins, macromolecules, and organelles. While autophagy has become one of the most attractive topics in cancer...

Mitogen-activated protein kinase-activated protein kinase-1 (MAPKAP-K1; also known as p90^rsk) contains two protein kinase domains in a single polypeptide. The N-terminal kinase domain is necessary for the phosphorylation of peptide substrates, whereas the C-terminal kinase domain is required for full activation of the N-terminal domain. Here we identify six sites in MAPKAP-K1a that become phosphorylated in transfected COS-1 cells. The inactive form of MAPKAP-K1a in unstimulated cells is partially phosphorylated at Ser²²² and Ser⁷³³. Stimulation with phorbol 12-myristate 13-acetate induces the phosphorylation of Thr³⁶⁰, Ser³⁶⁴, Thr⁵⁷⁴, and Ser³⁸¹ and increases the phosphorylation of Ser²²² and Ser⁷³³. Our data indicate that mitogen-activated protein kinase activates the C-terminal kinase domain by phosphorylating Thr⁵⁷⁴ and contributes to the activation of the N-terminal kinase domain by phosphorylating Ser³⁶⁴. The activated C-terminal domain phosphorylates Ser³⁸¹, which, together with phosphorylation of Ser³⁶⁴, activates the N-terminal kinase domain. The phosphorylation of Ser²²²and Ser⁷³³, which can be catalyzed by the N-terminal domain, does not activate MAPKAP-K1a per se, but Ser²²² phosphorylation appears to be required for activation. Ser²²², Ser³⁶⁴, and Ser³⁸¹ are situated in analogous positions to phosphorylation sites in protein kinase B, protein kinase C, and p70^S6K, suggesting a common mechanism of activation for these growth factor-stimulated protein kinases.

Mitogen-activated protein kinases (MAPKs) mediate cellular responses to a wide variety of extracellular stimuli. MAPK signal transduction cascades are tightly regulated, and individual MAPKs display exquisite specificity in recognition of their target substrates. All MAPK family members share a common phosphorylation site motif, raising questions as to how substrate specificity is achieved. Here we describe a peptide library screen to identify sequence requirements of the DEF site (docking site for ERK FXF), a docking motif separate from the phosphorylation site. We show that MAPK isoforms recognize DEF sites with unique sequences and identify two key residues on the MAPK that largely dictate sequence specificity. Based on these observations and computational docking studies, we propose a revised model for MAPK interaction with substrates containing DEF sites. Variations in DEF site sequence requirements provide one possible mechanism for encoding complex target specificity among MAPK isoforms.

EMBO Press is an editorially independent publishing platform for the development of EMBO scientific publications.