DIFFERENTIAL CDK-INHIBITOR GENE-EXPRESSION IN AGING HUMAN-DIPLOID FIBROBLASTS

Cellular aging is accompanied by a reduction in proliferative activity and changes in gene expression. To further elucidate the mRNA phenoty pe of aging fibroblasts we have monitored the expression of an array o f genes implicated in regulating cell-cycle progression, Fourteen gene s, including 3 cyclin-dependent kinase (CDK) inhibitors (p16I(INK4), p 21(SDI/CIP/WAF) and p27(KIP)), 5 cyclins, 4 CDKs, Cdi-1, and PCNA were tested in four primary fibroblast strains. Relative mRNA expression l evels were assessed using a rapid and sensitive Reverse Transcriptase- Polymerase Chain Reaction (RT-PCR) assay called the ''Primer-dropping' ' method. p16(INK4), a specific inhibitor of the cyclin D-associated k inases CDK4 and CDK6, was, in addition to p21 and cyclin D1, overexpre ssed in higher passage cells, while the abundance of the D-type kinase mRNAs remained relatively constant. Levels of cyclin H, a component o f the CDK-activating kinase (CAK) were markedly reduced in all strains examined, suggesting that the activity of target cyclin/CDK complexes may not be activated in aging cells. These results corroborate and ex tend previous observations demonstrating elevated expression of specif ic cell cycle genes in higher passage cells and suggest that overexpre ssion of the CDK-inhibitors p16(INK4) and p21(SDI)/(CIP/WAF), but not p27(KIP) may contribute to lower proliferative activity of senescing p rimary fibroblasts.