Murine FGF-inducible kinase is rapidly degraded via the nuclear ubiquitin-proteosome system when overexpressed in NIH 3T3 cells.
FGF-inducible kinase (Fnk) is a member of the polo-like kinase family of structurally-related serine/threonine protein kinases. These kinases appear to play critical roles in normal cell cycle progression and in the DNA damage response. In the case of Fnk, several reports indicate that this protein normally functions in cells as a stress-activated checkpoint kinase. However, when Fnk is ectopically overexpressed in cells, it likely becomes constitutively activated, and this promotes cell cycle arrest and apoptosis. In the present paper, we report that murine Fnk has a short half-life when transiently overexpressed in transfected NIH 3T3 fibroblasts. In contrast, when a kinase-deficient Fnk mutant protein, Fnk-K92M, is overexpressed in transfected cells, it is significantly more stable. We also found that Fnk-wild-type (WT) and Fnk-K92M are present in both the nucleus and cytoplasm of transfected cells and that Fnk nuclear export requires CRM1 function. Both of these proteins are degraded in cells via the nuclear ubiquitin-proteosome system; however, Fnk-K92M does not enter the nuclear compartment as efficiently as Fnk-WT and consequently it is significantly more stable. These results demonstrate that Fnk expression levels in transfected cells can be regulated by nuclear-cytoplasmic trafficking, ubiquitination, and proteosome-dependent degradation. Furthermore, our studies indicate that the downregulation of endogenous Fnk activity in stressed cells may occur, at least in part, by Fnk nuclear translocation and proteosomal degradation.
Alberts
Gregory F
GF
Department of Vascular Biology, Jerome H. Holland Laboratory for the Biomedical Sciences, American Red Cross, Rockville, Maryland 20855, USA.
Winkles
Jeffrey A
JA
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D051379:Mice; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D041681:NIH 3T3 Cells / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism*
D009097:Multienzyme Complexes; D025801:Ubiquitin; C093328:Plk3 protein, mouse; D017346:Protein Serine-Threonine Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex