Up-regulation of Survivin during Immortalization of Human Myofibroblasts Is Linked to Repression of Tumor Suppressor p16INK4a Protein and Confers Resistance to Oxidative Stress

Chin-Yi Kan,Carlotta Petti,Lauryn Bracken,Michelle Maritz,Ning Xu,Rosemary O'Brien,Chen Yang,Tao Liu,Jun Yuan,Richard B Lock,Karen L Mackenzie

doi:10.1074/jbc.m112.447821

Abstract

Survivin is an essential component of the chromosomal passenger complex and a member of the inhibitor of apoptosis family. It is expressed at high levels in a large variety of malignancies, where it has been implicated in drug resistance. It was also shown previously that survivin is up-regulated during telomerase-mediated immortalization, which occurs at a relatively early stage during carcinogenesis. This study shows that up-regulation of survivin during immortalization of human myofibroblasts is an indirect consequence of the repression of p16(INK4a). Survivin and p16(INK4a) were functionally linked by assays that showed that either the up-regulation of survivin or repression of p16(INK4a) rendered telomerase-transduced MRC-5 myofibroblasts resistant to oxidative stress. Conversely, siRNA-mediated down-regulation of survivin activated caspases and enhanced the sensitivity of immortal MRC-5 cells to oxidative stress. The E2F1 transcription factor, which is negatively regulated by the pRB/p16(INK4a) tumor suppressor pathway, was implicated in the up-regulation of survivin. Using the ChIP assay, it was shown that E2F1 directly interacted with the survivin gene (BIRC5) promoter in cells that spontaneously silenced p16(INK4a) during telomerase-mediated immortalization. E2F1 binding to the BIRC5 was also enhanced in telomerase-transduced cells subjected to shRNA-mediated repression of p16(INK4a). Together, these data show that repression of p16(INK4a) contributes to the up-regulation of survivin and thereby provides a survival advantage to cells exposed to oxidative stress during immortalization. The up-regulation of survivin during immortalization likely contributes to the vulnerability of immortal cells to transformation by oncogenes that alter intracellular redox state.

Highlights

Up-regulation of survivin and repression of the tumor suppressor p16INK4a often occur during cellular immortalization
It is shown that the up-regulation of survivin in human telomerase reverse transcriptase (hTERT)-immortalized myofibroblasts is intrinsically linked to repression of p16INK4a and underpins the resistance of immortal cells to oxidative stress, which may be advantageous during malignant transformation
Survivin Confers Resistance to Oxidant-induced Cell Death— Because survivin is up-regulated during telomerase-mediated immortalization of myofibroblasts (Fig. 2A, left panel, and Ref. 23), we investigated whether high levels of survivin contributed to the resistance of immortal MRC5hTERT-1 cells to oxidative stress

Summary

Background

Up-regulation of survivin and repression of the tumor suppressor p16INK4a often occur during cellular immortalization. E2F1 binding to the BIRC5 was enhanced in telomerase-transduced cells subjected to shRNA-mediated repression of p16INK4a Together, these data show that repression of p16INK4a contributes to the up-regulation of survivin and thereby provides a survival advantage to cells exposed to oxidative stress during immortalization. The very high frequency with which telomere maintenance mechanisms are activated and the p16INK4a/pRB pathway is disabled in human cancers attests to the relevance of these in vitro models of immortalization to the study of fundamental aspects of cancer cell biology [8, 9]. It is shown that the up-regulation of survivin in hTERT-immortalized myofibroblasts is intrinsically linked to repression of p16INK4a and underpins the resistance of immortal cells to oxidative stress, which may be advantageous during malignant transformation

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION