[en] Background: Telomerase confers cellular immortality by elongating telomeres, thereby circumventing the Hayflick limit. Extended-life-span cells have been generated by transfection with the human telomerase reverse transcriptase (hTERT) gene. hTERT transfected cell lines may be of outstanding interest to monitor the effect of drugs targeting the telomerase activity. The incidence of hTERT gene transfection at the proteome level is a prerequisite to that purpose. The effect of the transfection has been studied on the proteome of human fibroblast (W138). Cytosolic and nuclear fractions of W138 cells, empty vector transfected W138 (W138-HPV) and hTERT W138 cells were submitted to a 2D-DIGE (Two-Dimensional Differential In-Gel Electrophoresis) analysis. Only spots that had a similar abundance in W138 and W138-HPV, but were differentially expressed in W138 hTERT were selected for MS identification. This method directly points to the proteins linked with the hTERT expression. Number of false positive differentially expressed proteins has been excluded by using control W138-HPV cells. The proteome alteration induced by hTERT W138 transfection should be taken into account in subsequent use of the cell line for anti-telomerase drugs evaluation.
Results: 2D-DIGE experiment shows that 57 spots out of 2246 are significantly differentially expressed in the cytosolic fraction due to hTERT transfection, and 38 were confidently identified. In the nuclear fraction, 44 spots out of 2172 were selected in the differential proteome analysis, and 14 were identified. The results show that, in addition to elongating telomeres, hTERT gene transfection has other physiological roles, among which an enhanced ER capacity and a potent cell protection against apoptosis.
Conclusion: We show that the methodology reduces the complexity of the proteome analysis and highlights proteins implicated in other processes than telomere elongation. hTERT induced proteome changes suggest that telomerase expression enhances natural cell repair mechanisms and stress resistance probably required for long term resistance of immortalized cells. Thus, hTERT transfected cells can not be only consider as an immortal equivalent to parental cells but also as cells which are over-resistant to stresses. These findings are the prerequisite for any larger proteomics aiming to evaluate anti-telomerase drugs proteome alteration and thus therapeutics induced cell reactions.
Research Center/Unit :
Giga-Systems Biology and Chemical Biology - ULiège CART - Centre Interfacultaire d'Analyse des Résidus en Traces - ULiège
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Mazzucchelli, Gabriel ; Université de Liège - ULiège > Centre interfac. d'analyse des résidus en traces (CART)
Gabelica, Valérie ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Smargiasso, Nicolas ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Fleron, Maximilien ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Histologie - Cytologie
Ashimwe, Wilson; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Histologie - Cytologie
Rosu, Frédéric ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
De Pauw-Gillet, Marie-Claire ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Histologie - Cytologie - Département des sciences biomédicales et précliniques
Riou, Jean-François; CNRS, Museum Natl Hist Nat, INSERM,USM 503, U565,UMR 5153, Paris, France
De Pauw, Edwin ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Language :
English
Title :
Proteome alteration induced by hTERT transfection of human fibroblast cells
Publication date :
2008
Journal title :
Proteome Science
eISSN :
1477-5956
Publisher :
BioMed Central, United Kingdom
Volume :
6
Issue :
1
Pages :
12
Peer reviewed :
Peer Reviewed verified by ORBi
Funders :
F.R.S.-FNRS - Fonds de la Recherche Scientifique FRIA - Fonds pour la Formation à la Recherche dans l'Industrie et dans l'Agriculture
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