Abstract
Protein kinases carry out important functions in cells both by phosphorylating substrates and by means of regulated non-catalytic activities. Such non-catalytic functions have been ascribed to many kinases, including some members of the Ste20 family. The Drosophila Ste20 kinase Slik phosphorylates and activates Moesin in developing epithelial tissues to promote epithelial tissue integrity. It also functions non-catalytically to promote epithelial cell proliferation and tissue growth. We carried out a structure-function analysis to determine how these two distinct activities of Slik are controlled. We find that the conserved C-terminal coiled-coil domain of Slik, which is necessary and sufficient for apical localization of the kinase in epithelial cells, is not required for Moesin phosphorylation but is critical for the growth-promoting function of Slik. Slik is auto- and trans-phosphorylated in vivo. Phosphorylation of at least two of three conserved sites in the activation segment is required for both efficient catalytic activity and non-catalytic signaling. Slik function is thus dependent upon proper localization of the kinase via the C-terminal coiled-coil domain and activation via activation segment phosphorylation, which enhances both phosphorylation of substrates like Moesin and engagement of effectors of its non-catalytic growth-promoting activity.
Highlights
Slik kinase has catalytic activity-dependent and -independent functions
Summary of Slik phosphopeptides identified by LC-MS/MS
We were interested to know whether the regulatory mechanisms that control Slik catalytic activity, which is required for phosphorylation of the substrate Moesin and epithelial tissue integrity in vivo, control its non-catalytic growth-promoting function
Summary
Slik kinase has catalytic activity-dependent and -independent functions. Results: Mutation of activation segment phosphorylation sites abolishes both catalytic and non-catalytic activities; non-catalytic function depends upon localization via the C-terminal domain. Protein kinases carry out important functions in cells both by phosphorylating substrates and by means of regulated non-catalytic activities. Such non-catalytic functions have been ascribed to many kinases, including some members of the Ste[20] family. The Drosophila Ste[20] kinase Slik phosphorylates and activates Moesin in developing epithelial tissues to promote epithelial tissue integrity It functions non-catalytically to promote epithelial cell proliferation and tissue growth. Outside of the catalytic domain, which shows homology to the yeast kinase Ste20p, the 10 subfamilies show little sequence similarity to one another Despite their structural diversity, many Ste[20] kinases appear to regulate a few common cellular functions including cell proliferation and survival, cytoskeletal dynamics, and ion transport (2).
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