Phosphorylation and activation of nuclear Ca2+/calmodulin-dependent protein kinase phosphatase (CaMKP-N/PPM1E) by Ca2+/calmodulin-dependent protein kinase I (CaMKI)

► CaMKP-N/PPM1E underwent proteolytic processing and translocated to cytosol. ► The proteolysis was effectively inhibited by the proteasome inhibitors. ► Ser-480 of zebrafish CaMKP-N was phosphorylated by cytosolic CaMKI. ► Phosphorylation-mimic mutants of CaMKP-N showed enhanced activity. ► These r...

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Veröffentlicht in:Biochemical and biophysical research communications 2012-06, Vol.422 (4), p.703-709
Hauptverfasser: Onouchi, Takashi, Sueyoshi, Noriyuki, Ishida, Atsuhiko, Kameshita, Isamu
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Sprache:eng
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Zusammenfassung:► CaMKP-N/PPM1E underwent proteolytic processing and translocated to cytosol. ► The proteolysis was effectively inhibited by the proteasome inhibitors. ► Ser-480 of zebrafish CaMKP-N was phosphorylated by cytosolic CaMKI. ► Phosphorylation-mimic mutants of CaMKP-N showed enhanced activity. ► These results suggest that CaMKP-N is regulated by CaMKI. Nuclear Ca2+/calmodulin-dependent protein kinase phosphatase (CaMKP-N/PPM1E) is an enzyme that dephosphorylates and downregulates multifunctional Ca2+/calmodulin-dependent protein kinases (CaMKs) as well as AMP-dependent protein kinase. In our previous study, we found that zebrafish CaMKP-N (zCaMKP-N) underwent proteolytic processing and translocated to cytosol in a proteasome inhibitor-sensitive manner. In the present study, we found that zCaMKP-N is regulated by phosphorylation at Ser-480. When zCaMKP-N was incubated with the activated CaMKI, time-dependent phosphorylation of the enzyme was observed. This phosphorylation was significantly reduced when Ser-480 was replaced by Ala, suggesting that CaMKI phosphorylates Ser-480 of zCaMKP-N. Phosphorylation-mimic mutants, S480D and S480E, showed higher phosphatase activities than those of wild type and S480A mutant in solution-based phosphatase assay using various substrates. Furthermore, autophosphorylation of CaMKII after ionomycin treatment was more severely attenuated in Neuro2a cells when CaMKII was cotransfected with the phosphorylation-mimic mutant of zCaMKP-N than with the wild-type or non-phosphorylatable zCaMKP-N. These results strongly suggest that phosphorylation of zCaMKP-N at Ser-480 by CaMKI activates CaMKP-N catalytic activity and thereby downregulates multifunctional CaMKs in the cytosol.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2012.05.062