Difference between revisions of "Part:BBa K1699001"

Revision as of 13:52, 10 September 2015

Human short TERT promoter

Human short TERT promoter.

hTERT (human Telomerase Reverse Transcriptase) is a human promoter which controls the transcription of Telomerase, a gene highly expressed in cancer cells, and not in healthy ones (1).

Usage and Biology

hTERT promotes the transcription of the catalytic subunit of telomerase. Telomerase elongates the ends of chromosomes, regions called telomers. hTERT is not active in somatic cells and is highly active in most if not all cancer cell types. Therefore, the expression of an exogenous gene under the control of hTERT promoter will likely occur in cancer cells only. We have used this promoter as part of a two promoter system (the other one being survivin promoter, also highly active in cancer cells), in order drive the expression, and subsequently activate the core of our system (based on CRISPR?Cas9 technology) exclusively in cancer cells.

Characterization

The validity of hTERT promoter for the application of cancer-specific gene expression was performed using quantitative real-time PCR (qPCR), where the expression of hTERT was evaluated in several human cancer cell lines comprared to healthy cells (fibroblasts) (Fig. 1). The results show thousand-fold and higher TERT expression levels in cancer cells, suggesting marked promoter hyperactivation.

Fig. 1. hTERT epxression level in several human cell types, evaluated by qPCR.

This part was used by BGU 2015 team in a couple of constructs. Promoter activation worked in all of them.
1. hTERT GFP - robust transcription of GFP in all HepG2 cells. None detected in control (human fibroblasts).
2. hTERT dCas9-VP64 - activation system including gRNA under survivin promoter. Activation system worked in cancer cells and not in healthy ones.

References

1. The telomerase reverse transcriptase promoter drives efficacious tumor suicide gene therapy while preventing hepatotoxicity encountered with constitutive promoters http://www.nature.com/gt/journal/v8/n7/pdf/3301421a.pdf

Sequence and Features