Identification by suppression subtractive hybridization of p21 as a radio-inducible gene in human glioma cells.
BACKGROUND
Radio-gene therapy involves the delivery, to tumor cells, of a therapeutic transgene whose expression is controlled by irradiation.


METHODS
Here, we sought to identify novel radio-inducible transcripts in U87MG human malignant glioma cells using suppression subtractive hybridization (SSH).


RESULTS
Of 998 clones from a subtracted library of irradiated U87MG cells, 24 candidate clones were identified by dot blot and 3 clones were confirmed as having been induced by irradiation by Northern blot analysis. All three clones showed 99-100% homology to the cyclin-dependent kinase (cdk) inhibitor, p21(Waf/Cip1). A screening of 12 human malignant glioma cell lines revealed that irradiation increased p21 mRNA expression and p21 protein levels levels in all of the five cell lines retaining p53 wild-type activity in a p53 reporter assay, but in none of seven p53 reporter-negative cell lines.


CONCLUSIONS
Irradiation induces p21 mRNA expression in a strictly p53-dependent manner and may only enhance the expression of a limited number of genes in glioma cells. We conclude that the identification of radio-inducible genomic sequences suitable for radio-gene therapy may turn out to be difficult.
Rimner
A
A
Department of Neurology, University Tübingen, School of Medicine, Germany.
Wischhusen
J
J
Naumann
U
U
Gleichmann
M
M
Steinbach
J P
JP
Weller
M
M
D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000096:biosynthesis / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic / Q000528:radiation effects*; D015316:Genetic Therapy / Q000379:methods*; D005910:Glioma / Q000235:genetics* / Q000628:therapy; D006801:Humans; D009693:Nucleic Acid Hybridization; D014407:Tumor Cells, Cultured
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins