- HPV-16 E6/E7 Gene Expression Constructs
- SV40 T Antigen Expression Constructs
- Telomerase Expression Constructs
- Tumor Suppression Gene Inactivation Constructs
A way around this is the use of immortalized cells, which have an extended replicative capacity and enable you to use the same consistent cells throughout your research project.
Several methods exist for immortalizing mammalian cells in culture conditions.
With years of experience in cell immortalization, our partner ABM have developed the most comprehensive cell immortalization product line comprising of recombinant lentiviral, retroviral (MMLV) and adenoviral viruses expressing EBV, HPV-16 E6/7 and SV40 T antigens, hTERT, p53 and RB siRNAs, and ras & myc mutants. All these tools will make your cell immortalization project simpler and easier than ever before.
Virus Induction: EBV, HPV E6/7 and SV40 T Antigen
One method is to use viral genes, such as EBV, HPV-16 E6/7 gene, and the Simian Virus 40 (SV40) T antigens, to induce immortalization. SV40 T antigens have been shown to be the simplest and most reliable agent for the transformation of many different cell types in culture and the mechanism of SV40 T antigens in cell immortalization is well studied. For the most part, viral genes achieve immortalization by inactivating the tumor suppressor genes (p53, Rb, and others) that can induce a replicative senescent state in cells. Recent studies have also shown that SV40 T antigen can induce Telomerase activity in the infected cells.
A more recently discovered approach to cell immortalization is through the expression of Telomerase Reverse Transcriptase protein (TERT), particularly for cells that are most affected by telomere length (e.g., human). This protein is inactive in most somatic cells, but when hTERT is exogenously expressed, the cells are able to maintain sufficient telomere lengths to avoid replicative senescence. Analysis of several telomerase-immortalized cell lines has verified that the cells maintain a stable genotype and retain critical phenotypic markers.
However, over-expression of hTERT in some cell types (especially in primary epithelial cells) fails to induce cell immortalization and it may induce cell death.
Inactivation of Tumor Suppression Genes
Recent research has found that co-expression of hTERT catalytic subunit with either p53 or RB siRNA can immortalize human primary ovarian epithelial cells, providing a more authentic normal cell model with well-defined genetic background (Yang G. et al. , Carcinogenesis 2007; Yang G. et al ., Oncogene 2007) . Likewise, overexpression of Ras or Myc T58A mutants have also been found to be able to immortalize some primary cell types (Sears R. et al ., Genes & Dev. 2000). The HOX genes (HOXB8, HOXA9, and HOXA10) are a family of homeodomain-containing transcription factors associated with malignancies such as acute myeloid leukemia and acute lymphoid leukemia. Accordingly, overexpression of the HOX genes can be used to immortalize various hematopoietic cells, including macrophages, hematopoietic progenitor cells, and myeloid progenitor cells.