Protein kinase C epsilon regulation of brain-specific serine/threonine-protein kinase 1 kinase activity and nuclear localization



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Protein Kinase C epsilon (PKCε) signaling plays a role in a variety of physiological functions, including neurotransmission, cell cycle progression, and responses to ethanol. However, only a few downstream substrates of PKCε that contribute to these functions are known. Investigating PKCε substrates can further reveal the role of PKCε in these processes and provide targets for the treatment of conditions associated with the function of PKCε. Recently, a chemical genetic screen identified brain-specific serine/threonine-protein kinase 1 (BRSK1) as a substrate of PKCε, with phospho-sites at serine 555 (S555) and serine 559 (S559). BRSK1 plays a role in regulating proper neuronal development and neurotransmitter release in mature neurons. Currently, it is not known if PKCε phosphorylation of BRSK1 at these sites regulates BRSK1 activity or cellular function. In this study, we used phospho-mimic and phospho-null mutations at both S555 and S559, as well as in vitro kinase assays to determine the effect of phosphorylation at these sites on BRSK1 kinase activity. Additionally, we used immunofluorescence to determine if these mutations or if pharmacological activation or inhibition of PKCε changes the nuclear localization of BRSK1 in Neuro-2a cells. We found that phospho-mimic and phospho-null mutations at S555 and S559 as well as preincubation of BRSK1 with PKCε resulted in decreased BRSK1 kinase activity. However, these manipulations did not alter BRSK1 localization. These results suggest that PKCε phosphorylation of BRSK1 decreases BRSK1 kinase activity.



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