Reduction in mortalin level by its antisense expression causes senescence-like growth arrest in human immortalized cells

Abstract

Background: Human normal cells have active p53 and pRB tumor suppressor pathways and undergo telomere shortening at each cell division. When immortalized, these exhibit functional inactivation of one or both tumor suppressor pathways and activation of telomere-maintaining mechanisms. Regulation of immortalization promoting molecular pathways by other genes is poorly understood and is an essential component of cancer therapeutics. In the present study, we have immortalized human normal cells by functional inactivation of p53 and pRB tumor suppressor proteins and simultaneous activation of telomerase function. We demonstrate that when the expression of mortalin (a member of hsp70 family proteins) was suppressed in such genetically defined immortal cells they underwent a permanent growth arrest.

Methods: WI-38 cells were transfected with expression plasmids encoding papilloma virus E6 and E7 proteins and a catalytic subunit of human telomerase enzyme. The derivative cells were compromised for p53 and pRB senescence pathways, showed telomerase activity and were immortalized. These cells were transfected with antisense expression plasmid for mortalin. The derivative clones were analyzed for mortalin expression, proliferation and telomerase activity.

Results: WI-38 cells were immortalized with E6, E7 and the catalytic subunit of human telomerase. The derivative-immortalized cells when suppressed for mortalin expression underwent senescence-like growth arrest.

Conclusions: The data demonstrate that suppression of mortalin is sufficient to induce growth arrest in human immortalized cells that have compromised p53 and pRB functions and possess telomerase activity.