DNA damage and repair in lymphoblastoid cell lines from normal donors and fragile X syndrome patients.
BACKGROUND
Because lymphocytes from fragile X patients have been reported as hypersensitive to bleomycin-induced chromatid breaks and because the number of trinucleotide repeats in families with fragile X syndrome has a propensity to expand, we have investigated the possibility that fragile X cells may be hypersensitive to DNA damage and have a lower capacity for DNA repair.


METHODS
Lymphocytes from normal and fragile X syndrome donors were immortalized by Epstein-Barr virus transformation. Characteristics of fragile X syndrome including the folate-sensitive fragile site on chromosome Xq27.3, length of CGG repeat expansion, and FMRP expression in Epstein-Barr virus-transformed lymphoblastoid cell lines were analyzed by standard cytogenetic methods, Southern blot, and Western blot, respectively. Analysis of DNA damage and repair induced by hydrogen peroxide, bleomycin, ethyl methanesulfonate, 4-nitroquinoline-N-oxide, etoposide, and mitomycin C was carried out by single-cell gel electrophoresis assay (known as comet assay).


RESULTS
Lymphoblastoid cell lines from fragile X donors had a folate-sensitive fragile site on chromosome Xq27.3, no or low FMRP expression, and expansion of the CGG repeat. Results of comet assay showed that fragile X cells were not more sensitive to mutagen-induced DNA strand breaks and did not have lower DNA repair capacity in comparison with normal cells. Furthermore, one fragile X cell line showed hyposensitivity to DNA strand breaks induced by hydrogen peroxide, bleomycin, and ethyl methansulfonate.


CONCLUSIONS
The results of this study do not support the notion that CGG trinucleotide expansion in fragile X syndrome is caused by permanent deficiency in DNA repair.
Wang
Tsu Shing
TS
Department of Life Sciences, Chung Shan Medical University, Taichung 402, Taiwan, Republic of China (ROC). tsushing@csmu.edu.tw
Hsieh
Lei Jiau
LJ
Hsu
Tsung Yang
TY
Chung
Chiao Hui
CH
Li
Shuan Yow
SY
D015112:4-Nitroquinoline-1-oxide / Q000494:pharmacology; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D001761:Bleomycin / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002460:Cell Line; D002472:Cell Transformation, Viral; D020552:Comet Assay; D004249:DNA Damage; D004260:DNA Repair; D005020:Ethyl Methanesulfonate / Q000494:pharmacology; D005047:Etoposide / Q000494:pharmacology; D005260:Female; D051860:Fragile X Messenger Ribonucleoprotein 1; D005600:Fragile X Syndrome / Q000235:genetics*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008214:Lymphocytes / Q000187:drug effects / Q000502:physiology*; D008297:Male; D016685:Mitomycin / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000235:genetics; D016877:Oxidants / Q000494:pharmacology; D016601:RNA-Binding Proteins; D019680:Trinucleotide Repeat Expansion / Q000235:genetics
D015112:4-Nitroquinoline-1-oxide; D000903:Antibiotics, Antineoplastic; D018906:Antineoplastic Agents, Alkylating; D000972:Antineoplastic Agents, Phytogenic; D001761:Bleomycin; D002273:Carcinogens; D005020:Ethyl Methanesulfonate; D005047:Etoposide; D051860:Fragile X Messenger Ribonucleoprotein 1; D006861:Hydrogen Peroxide; D016685:Mitomycin; D009419:Nerve Tissue Proteins; D016877:Oxidants; D016601:RNA-Binding Proteins; C497683:FMR1 protein, human