Over expression of the selectable marker blasticidin S deaminase gene is toxic to human keratinocytes and murine BALB/MK cells.
BACKGROUND
The blasticidin S resistance gene (bsr) is a selectable marker used for gene transfer experiments. The bsr gene encodes for blasticidin S (BS) deaminase, which has a specific activity upon BS. Therefore, its expression is supposed to be harmless in cells. The work reported on herein consisted of experiments to verify a possible toxicity of bsr on mammalian cells, which include several cell lines and primary cultures.


RESULTS
Murine keratinocyte BALB/MK and human primary keratinocyte cells transduced with the retroviral vector LBmSN, which has an improved expression system of bsr, namely bsrm, died in five days after the transduction. Meanwhile the control vector LBSN, which expresses bsr, did not provoke cell death. The lethal activity of bsrm was observed only in human keratinocytes and BALB/MK cells among the cell types tested here. Death appears to be mediated by a factor, which is secreted by the BALB/MK transduced cells.


CONCLUSIONS
By our study we demonstrated that the expression of bsrm gene is toxic to human keratinocytes and BALB/MK cells. It is likely over expression of BS deaminase gene is responsible for the death.
Bento
Fernanda Mara
FM
Department of Biophysics & Center for Gene Therapy, UNIFESP - EPM, São Paulo, SP, Brazil. fmbento@hotmail.com <fmbento@hotmail.com>
Takeshita
Daniela
D
Sacramento
Chester Bittencourt
CB
Machado
Tamara Rocha
TR
Mathor
Monica Beatriz
MB
Carmona
Adriana Karaoglanovic
AK
Han
Sang Won
SW
D000619:Aminohydrolases / Q000235:genetics* / Q000633:toxicity*; D000818:Animals; D002452:Cell Count; D016923:Cell Death / Q000235:genetics; D002460:Cell Line; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D005804:Genes, Lethal / Q000235:genetics; D005819:Genetic Markers / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics / Q000633:toxicity; D006801:Humans; D015603:Keratinocytes / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D041681:NIH 3T3 Cells / Q000737:chemistry / Q000378:metabolism; D009705:Nucleosides / Q000235:genetics / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods
D005819:Genetic Markers; D009705:Nucleosides; C004500:blasticidin S; D000619:Aminohydrolases; C019795:blasticidin S deaminase