Extension of Life-Span by Introduction of Telomerase into Normal Human Cells

Andrea G.Bodnar et al.

Science,pp. 349-352,

vol. 279 (1998)

Podlevsky, J.D., Bley, C.J., Omana, R.V., Qi, X., Chen, J. (2007) The Telomerase Database. Nucleic Acids Res. 36 D339-D343.

Telomeres senescence theory

Telomeres senescence theory

Human telomeres (in yellow)

Telomeres senescence theory

Human telomeres (in yellow)

- TTAGGG/CCCTAA sequence- Synthesized by theribonucleoproteinenzyme telomerase- Active in germline cells, keeps telomeres at about 15 kpb

- Not expressed in most human somatic tissues, where telomeres lenghts is significantly shorter

Telomeres senescence theory

Human telomeres (in yellow)

- TTAGGG/CCCTAA sequence- Synthesized by theribonucleoproteinenzyme telomerase- Active in germline cells, keeps telomeres at about 15 kpb

- Not expressed in most human somatic tissues, where telomeres lenghts is significantly shorter

Theory proposes that cells become senescent when progressive telomeres shortening during

each division produces a treshold telomere length

How telomerase works

How telomerase works

How telomerase works

How telomerase works

How telomerase works

Research...

Research...- The hTRT has been cloned- telomerase activity can be reconstituted by transient expression of hTRT in norman human diploid cells- hTR (template RNA component costituvely expressed)

- hTRT- normal cells transfected with2 different hTRT expression constructs:

Research...- The hTRT has been cloned- telomerase activity can be reconstituted by transient expression of hTRT in norman human diploid cells- hTR (template RNA component costituvely expressed)

- hTRT- normal cells transfected with2 different hTRT expression constructs:

#1engineered by removal of the 5’ and 3’ untranslatedregions of hTRT and creation of Kozak consensus sequence

Research...- The hTRT has been cloned- telomerase activity can be reconstituted by transient expression of hTRT in norman human diploid cells- hTR (template RNA component costituvely expressed)

- hTRT- normal cells transfected with2 different hTRT expression constructs:

#1#2

engineered by removal of the 5’ and 3’ untranslatedregions of hTRT and creation of Kozak consensus sequence

native sequence cloned downstream of the SV40 promoter

Research...- The hTRT has been cloned- telomerase activity can be reconstituted by transient expression of hTRT in norman human diploid cells- hTR (template RNA component costituvely expressed)

- hTRT- normal cells transfected with2 different hTRT expression constructs:

#1#2

comparison between life span of MPSV-hTRTtransfected cells and vector only transfected cells

comparison of lifespan of activity positive and activity negative stable clones containing SV40-hTRT constructs

Telomerase activity in stable Retinal Pigment Epithelial cells

Telomerase activity in stable Retinal Pigment Epithelial cells

Telomeres length in stable RPE and BJ clones

Telomeres length in stable RPE and BJ clones

Effects of telomerase expression on cell lifespan

Effects of telomerase expression on cell lifespan

Effects of telomerase expression on cell lifespan

Effects of telomerase expression on cell lifespan

Implications

ImplicationsResults indicate that telomere loss in the absence of

telomerase is the intrinsic timing mechanism that controls the number of cell divisions prior to senescence

Very low level of telomerase activity are apparently insufficient to prevent telomere shortening.

This is consistent with the observation that stem cells have low but detectable telomerase activity, yet continue to exhibit shortening of their telomeres throughout life

Promoter strength, structure of untranslated regions, site of integration, levels of hTR and hTRT and telomere- or

telomerase-associated proteins in specific cell types are all factors that may affect the functional level of telomerase

Ideas

Ideas- against atrophy of skin through loss of extracellular matrix

homeostasis in dermal fibroblasts

- age-related macular degeneration of lipofuscin and down-regulation of a neuronal survival factor in RPE cells

- atherosclerosis caused by loss of proliferative capacity and overexpression of hypertensive and thrombotic factors in

endothelial cells

- replacing of tumor cell lines with cloned normal diploid cells

- production of normal or engineered biotechnology products or gene therapy

Thank you for listening

Original paper DOI: 10.1126/science.279.5349.349

pdf version available at http://www.slideshare.net/ATMB