CDK2 translational down-regulation during endothelial senescence.
Here we report for the first time that loss of CDK2 activity, by translational inhibition and through CDK2 inhibition by p21(Cip1/Waf1), may be responsible for endothelial senescence. We show that expression of dominant-negative p53 extends human umbilical vein endothelial cell (HUVEC) lifespan past senescence. HUVEC expressing telomerase can completely bypass senescence and become immortal (i-HUVEC). Surprisingly, early passage i-HUVEC, like senescent HUVEC, express high levels of the CDK inhibitors p16(INK4a) and p21(Cip1/Waf1). Expression of p16(INK4a) can persist for over 280 population doublings, while p21(Cip1/Waf1) expression was eventually lost in five of six i-HUVEC lines. Senescent HUVEC contain undetectable CDK2 activity, which results from a dramatic reduction of CDK2 protein levels and inhibition of remaining CDK2 by p21(Cip1/Waf1). The decreased CDK2 levels in senescent HUVEC are not due to decreased transcription or protein stability; rather, CDK2 translation declines during senescence. Bypass of endothelial senescence by telomerase entails the restoration of CDK2 translation and activity. These results suggest that p16(INK4a) does not play a role in endothelial senescence. Rather, CDK2 translational down-regulation may be a key regulatory event in replicative senescence of endothelial cells. Understanding the mechanisms regulating endothelial senescence will be critical in determining the role of endothelial senescence in tumor growth.
Freedman
Deborah A
DA
Vascular Biology Program, Department of Surgery, Children's Hospital, 1 Blackfan Circle, Harvard University Medical School, Karp Family Research Laboratories, Floor 12, Boston, MA 02115, USA.
Folkman
Judah
J
D015153:Blotting, Western; D042846:CDC2-CDC28 Kinases / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D018929:Cell Culture Techniques; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D016922:Cellular Senescence; D002999:Clone Cells; D051357:Cyclin-Dependent Kinase 2; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000037:antagonists & inhibitors; D050759:Cyclin-Dependent Kinase Inhibitor p21; D015536:Down-Regulation; D004730:Endothelium, Vascular / Q000166:cytology; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008938:Mitosis; D008954:Models, Biological; D011233:Precipitin Tests; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019098:Telomerase / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D049452:Green Fluorescent Proteins; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D019098:Telomerase