Epstein-Barr virus transformation of human lymphoblastoid cells from patients with fragile X syndrome induces variable changes on CGG repeats size and promoter methylation.
BACKGROUND
Our understanding of fragile X syndrome can be improved by reversing the expression of the silenced fragile X mental retardation 1 (FMR1) gene in immortalized cells from these patients. Epstein-Barr virus (EBV) infection has been extensively used to transform B cells into a permanent lymphoblastoid cell line.


METHODS
We immortalized B lymphocytes from three different fragile X patients and one normal male. We analyzed the CGG triplet repeats and methylation status of the FMR1 and interferon (IFN)-gamma promoter. We also assayed FMR1 mRNA levels by real-time PCR and FMR1 protein (FMRP) by Western blot.


RESULTS
We observed that EBV transformation may induce the instability of CGG repeats and DNA demethylation that can lead to the modification of mRNA expression.


CONCLUSIONS
EBV transformation may induce variable changes in the genome that can lead to the misinterpretations of experimental data obtained from these cells. Thus, periodic testing of DNA from immortalized cells should be routinely undertaken to detect undesired effects.
Bonilla
Victoria
V
Department of Biochemical Medicine and Molecular Biology, Faculty of Medicine and University Hospital of the Virgin Macarena, University of Seville, Seville, Spain.
Sobrino
Francisco
F
Lucas
Miguel
M
Pintado
Elizabeth
E
D001483:Base Sequence; D015139:Blotting, Southern; D002460:Cell Line; D002472:Cell Transformation, Viral; D017124:Conserved Sequence; D019175:DNA Methylation; D015226:Dinucleoside Phosphates / Q000235:genetics; D005600:Fragile X Syndrome / Q000235:genetics* / Q000821:virology*; D004854:Herpesvirus 4, Human / Q000472:pathogenicity*; D006801:Humans; D008607:Intellectual Disability / Q000235:genetics*; D007360:Intelligence; D008214:Lymphocytes / Q000502:physiology / Q000821:virology; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D018911:Trinucleotide Repeats / Q000235:genetics*
D015226:Dinucleoside Phosphates; C015772:CpG dinucleotide