Molecular mechanisms of impaired stimulation of DNA synthesis in cultured hepatocytes of aged rats.
We examined epidermal growth factor (EGF)- and epinephrine-stimulated mitogen-activated protein kinase kinase (MEK) 1 and MEK2 activities, DNA polymerase alpha activity, and EGF-stimulated E2F DNA binding activity in primary cultured hepatocytes from 6- and 24-mo-old rats. MEK stimulation by either EGF or epinephrine was not altered with aging. However, stimulation of DNA polymerase alpha activity by these agents was 70% and 50% lower, respectively, in cells of aged compared with cells of young rats, consistent with a lesser increase in [3H]thymidine incorporation. EGF-stimulated E2F (a transcription factor that regulates expression of the DNA polymerase alpha gene) binding to DNA was reduced with age. PD-098059, a specific inhibitor of MEK, inhibited EGF-stimulated MEK1 and MEK2 activities in hepatocytes from 6- and 24-mo-old rats. Although PD-098059 inhibited EGF-stimulated DNA synthesis in hepatocytes from 6-mo-old rats, it had no effect in 24-mo-old rats. Thus the age-related impairment appears to occur before E2F activation, and signal transduction sequences other than the mitogen-activated protein kinase pathway may be involved in stimulated DNA synthesis in hepatocytes from old rats.
Kitano
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S
Molecular Physiology and Genetics Section, Laboratory of Cellular and Molecular Biology, Gerontology Research Center, Baltimore, Maryland 21224, USA.
Fawcett
T W
TW
Yo
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Y
Roth
G S
GS
D000375:Aging / Q000502:physiology*; D000818:Animals; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis*; D004256:DNA Polymerase I / Q000378:metabolism; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D004815:Epidermal Growth Factor / Q000494:pharmacology; D004837:Epinephrine / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism*; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D008297:Male; D020929:Mitogen-Activated Protein Kinase Kinases; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D056465:Retinoblastoma-Binding Protein 1; D013936:Thymidine / Q000378:metabolism; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000378:metabolism
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; D004815:Epidermal Growth Factor; D004247:DNA; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D020929:Mitogen-Activated Protein Kinase Kinases; D004256:DNA Polymerase I; D013936:Thymidine; D004837:Epinephrine