Role of the regulatory domain of protein kinase D2 in phorbol ester binding, catalytic activity, and nucleocytoplasmic shuttling.
Protein kinase D2 (PKD2) belongs to the PKD family of serine/threonine kinases that is activated by phorbol esters and G protein-coupled receptors (GPCRs). Its C-terminal regulatory domain comprises two cysteine-rich domains (C1a/C1b) followed by a pleckstrin homology (PH) domain. Here, we examined the role of the regulatory domain in PKD2 phorbol ester binding, catalytic activity, and subcellular localization: The PH domain is a negative regulator of kinase activity. C1a/C1b, in particular C1b, is required for phorbol ester binding and gastrin-stimulated PKD2 activation, but it has no inhibitory effect on the catalytic activity. Gastrin triggers nuclear accumulation of PKD2 in living AGS-B cancer cells. C1a/C1b, not the PH domain, plays a complex role in the regulation of nucleocytoplasmic shuttling: We identified a nuclear localization sequence in the linker region between C1a and C1b and a nuclear export signal in the C1a domain. In conclusion, our results define the critical components of the PKD2 regulatory domain controlling phorbol ester binding, catalytic activity, and nucleocytoplasmic shuttling and reveal marked differences to the regulatory properties of this domain in PKD1. These findings could explain functional differences between PKD isoforms and point to a functional role of PKD2 in the nucleus upon activation by GPCRs.
Department of Internal Medicine l, Medical University of Ulm, Ulm 89081, Germany.
Alexandra
A
Auer
Julia
J
von Blume
Sabine
S
Sturany
Götz
G
von Wichert
Johan
J
Van Lint
Jackie
J
Vandenheede
Guido
G
Adler
Thomas
T
Seufferlein
D002384:Catalysis; D002460:Cell Line; D045744:Cell Line, Tumor; D006801:Humans; D028884:Karyopherins / Q000502:physiology; D029543:Nucleocytoplasmic Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D010703:Phorbol Esters / Q000378:metabolism*; D011493:Protein Kinase C / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D000091283:Protein Kinase D2; D011494:Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D021382:Protein Sorting Signals / Q000502:physiology; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D018160:Receptors, Cytoplasmic and Nuclear / Q000502:physiology; D013274:Stomach Neoplasms / Q000201:enzymology; D014316:Tritium; D000097606:Exportin 1 Protein
D028884:Karyopherins; D029543:Nucleocytoplasmic Transport Proteins; D015240:Phorbol 12,13-Dibutyrate; D010703:Phorbol Esters; D011493:Protein Kinase C; D000091283:Protein Kinase D2; D011494:Protein Kinases; D021382:Protein Sorting Signals; D018160:Receptors, Cytoplasmic and Nuclear; D014316:Tritium; D000097606:Exportin 1 Protein; C089212:protein kinase D