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Telomerase is a specialized reverse transcriptase responsible for synthesizing telomeric DNA at the chromosome ends. We have previously found that all the six telomerase subunit proteins (hTERT, hTR, TEP1, hsp90, p23 and dyskerin) are needed for the full enzyme activity. Telomerase activity has been reported up-regulated by PKC but the detail mechanism is not clear. In this study, we examine how PKC regulates telomerase activity in oral cancer cells. Exposure cells to a PKC inhibitor, binsindolylmaleimide I (BIS), resulted in the inhibition of telomerase activity in a dose-dependent manner, but the mRNA levels of telomerase subunits were not influenced, suggesting a post-translational regulatory mechanism may exist. RNA interference studies reveal that inhibition of PKC isoforms α, β, δ, ε, ζ specifically reduced telomerase activity. In vitro phosphorylations by PKC α, βI, βII, δ, ε, ζ can restore BIS-inhibited telomerase activity, and the phosphorylation target is on hTERT, suggesting hTERT phosphorylation by the PKC isoenzyme is crucial for telomerase activity. Treatment of hsp-90 inhibitor, novobiocin (C-terminal inhibitor) can dissociate hsp90-hTERT association as revealed by immunoprecipitation and immunoblot analysis along with the reduction of telomerase activity. However, further treatment of PKC activator, SC-10 can restore the association of hsp90 and hTERT and reactivate telomerase, suggesting hTERT phosphorylation by PKC is essential for telomerase holoenzyme integrity and function. In conclusion, PKC α, βI, βII, δ, ε, ζ can phosphorylate hTERT in oral cancer cells. This phosphorylation is essential for telomerase holoenzyme assembly, and the structure integrity is necessary for telomerase full enzyme activity.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]