Regulation of Ack1 localization and activity by the amino-terminal SAM domain

<p>Abstract</p> <p>Background</p> <p>The mechanisms that regulate the activity of the nonreceptor tyrosine kinase Ack1 (activated Cdc42-associated kinase) are poorly understood. The amino-terminal region of Ack1 is predicted to contain a sterile alpha motif (SAM) domain. SAM domains share a common fold and mediate protein-protein interactions in a wide variety of proteins. Here, we addressed the importance of the Ack1 SAM domain in kinase activity.</p> <p>Results</p> <p>We used immunofluorescence and Western blotting to show that Ack1 deletion mutants lacking the N-terminus displayed significantly reduced autophosphorylation in cells. A minimal construct comprising the N-terminus and kinase domain (NKD) was autophosphorylated, while the kinase domain alone (KD) was not. When expressed in mammalian cells, NKD localized to the plasma membrane, while KD showed a more diffuse cytosolic localization. Co-immunoprecipitation experiments showed a stronger interaction between full length Ack1 and NKD than between full length Ack1 and KD, indicating that the N-terminus was important for Ack1 dimerization. Increasing the local concentration of purified Ack1 kinase domain at the surface of lipid vesicles stimulated autophosphorylation and catalytic activity, consistent with a requirement for dimerization and trans-phosphorylation for activity.</p> <p>Conclusions</p> <p>Collectively, the data suggest that the N-terminus of Ack1 promotes membrane localization and dimerization to allow for autophosphorylation.</p

Additional file 1: of A comparative analysis of vaccine administration in urban and non-urban skilled nursing facilities

Customized database used in this study. (XLSX 2173 kb