NPM/ALK downregulates p27Kip1 in a PI-3K-dependent manner.
OBJECTIVE
Anaplastic large-cell lymphomas (ALCL) are frequently associated with the chromosomal translocation t(2;5) (p23;q35) resulting in the NPM/ALK fusion gene that encodes a constitutively activated tyrosine kinase. We showed that NPM/ALK stimulated cell proliferation and that PI-3K/AKT pathway played an important role in this effect. p27Kip1 is a member of the CDK family inhibitory proteins regulating the entry into S phase. It was reported that p27Kip1 function is impaired in many tumors. In this study we investigated the role of PI-3K/AKT in NPM/ALK-dependent downregulation of p27Kip1 protein.


METHODS
To investigate this phenomenon the pro-B cell line BaF3, BaF3 cell line stably expressing NPM/ALK, and ALCL SUP-M2 cell line were used. The p27Kip1 protein expression before and after LY294002, wortmannin, or epoxomicin treatment and phosphorylation status of AKT were measured in parental and NPM/ALK+ cells by Western analysis. Also, the localization of p27Kip1 protein was analyzed by fractionation and immunoblotting.


RESULTS
p27Kip1 was found to be downregulated in NPM/ALK-transformed hematopoietic cells, but inhibition of proteasome-dependent degradation pathway by epoxomicin reversed this effect. In addition, treatment of NPM/ALK+ cells with LY294002, the PI-3K inhibitor, caused elevation of p27Kip1 protein expression and its nuclear localization.


CONCLUSIONS
Taken together, we postulate that NPM/ALK-PI-3K pathway stimulates cell proliferation by regulation of the expression and nuclear localization of p27Kip1.
Slupianek
Artur
A
College of Science and Technology, Center for Biotechnology, Temple University, Philadelphia, Pa. 19008, USA. aslupian@temple.edu
Skorski
Tomasz
T
D001402:B-Lymphocytes / Q000166:cytology / Q000378:metabolism / Q000473:pathology; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D002867:Chromones / Q000494:pharmacology; D050760:Cyclin-Dependent Kinase Inhibitor p27; D015536:Down-Regulation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects; D006412:Hematopoietic Stem Cells / Q000378:metabolism / Q000473:pathology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D016403:Lymphoma, Large B-Cell, Diffuse / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D009025:Morpholines / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D016196:S Phase / Q000187:drug effects / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D014170:Transformation, Genetic; D014178:Translocation, Genetic / Q000235:genetics
C436291:CDKN1B protein, human; D002352:Carrier Proteins; D002867:Chromones; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D009025:Morpholines; D009842:Oligopeptides; D050760:Cyclin-Dependent Kinase Inhibitor p27; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; C107023:p80(NPM-ALK) protein; D011505:Protein-Tyrosine Kinases; D046988:Proteasome Endopeptidase Complex; C078846:epoxomicin