Regulation of hTERT expression and NFkappaB signaling by the HPV16 E6 oncoprotein

Understanding the mechanism by which high risk human papillomaviruses (HR HPV) induce telomerase activity is important for cancer research because hTERT, the catalytic subunit of telomerase, is critical for cell immortalization and transformation. The HR HPV E6 oncoprotein utilizes multiple mechanisms to induce telomerase. Importantly, multiple transcription factors are found to be involved in the regulation of hTERT expression by E6.;NFX1-91 functioned as a repressor of hTERT in keratinocytes through interacting with the inhibitory mSin3A-HDAC complex. Degradation of NFX1-91 by E6/E6AP or knockdown of NFX1-91 by siRNA released the mSin3A-HDAC complex, which allowed histone acetylase recruitment to the hTERT promoter, and induction of hTERT expression. c-Myc binding to E boxes at the hTERT promoter was necessary but not sufficient for hTERT expression. E6 expression resulted in GCN5 recruitment to the hTERT promoter, which was E box dependent, suggesting c-Myc's role in regulating histone acetylation. Another transcription factor, MAZ was also found at the hTERT promoter and functioned to repress hTERT expression. E6 reduced MAZ binding, and correspondingly increased Sp1 binding at the hTERT promoter, which was shown previously to synergistically induce hTERT expression with c-Myc. Overall, E6 coordinates multiple transcriptional activators and repressors to activate hTERT expression.;In addition to regulation of hTERT expression, other downstream targets of NFX1-91 might have a role in regulation of the HPV life cycle. Microarray analysis revealed that knockdown of NFX1-91 induced upregulation of NFkappaB-responsive genes, by altering NFkappaB signaling. Downregulation of NFkappaB inhibitors (p105) was observed in cells in which NFX1-91 was knocked down. NFX1-91 was shown to bind to the p105 promoter and upregulate p105 expression. Degradation of NFX1-91 by E6 expression or knockdown of NFX1-91 by siRNA reduced p105 expression, and upregulated NFkappaB targets (IAPs, Bcls, TNF and multiple cytokines and their receptors). These data suggest a mechanism by which HPV E6 activates the NFkappaB pathway by degrading NFX1-91.