The effect of bisphosphonates on gene expression: GAPDH as a housekeeping or a new target gene?

We evaluated GAPDH gene expression by real time RT PCR in breast (MCF-7 and T47D) and prostate (PC3 and DU-145) cancer cell lines treated with amino and non-amino bisphosphonates.Our results showed that amino-bisphosphonates significantly decrease in a dose-dependent manner the expression of GAPDH gene.Therefore, GAPDH is inaccurate to normalize mRNA levels in studies investigating the effect of bisphosphonates on gene expression and it should be avoided. On the other hand, this gene could be considered a potential target to observe the effects of bisphosphonates on cancer cells.GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) is well known for its glycolytic function of converting D-Glyceraldeide-3-phosphate to 1,3-bisphosphoglycerate and it has been commonly considered as a constitutive housekeeping gene. It is widely used as a control RNA in Northern Blotting and in RT-PCR analysis and recently in real time RT-PCR.In some experimental systems its expression is constant at different times and after experimental manipulation [1]. In breast cancer cells treated with endoxifen GAPDH was used to normalize the expression data of the progesterone receptor mRNA [2]. In addition, GAPDH was the best control gene in the apoptosis pattern on the myeloid cell lines incubated with Camptothecin investigated by real time RT-PCR [3].However, there is overwhelming evidence suggesting that its use as an internal standard is inappropriate [4]. Growth hormone, oxidative stress and the tumour suppressor TP53 have all been shown to activate its transcription, which can also be induced in endothelial cells [5]. Conversely, retinoic acid down-regulates GAPDH transcription in adipocytes [6]. Furthermore, it has been observed that GAPDH mRNA expression was not normal in some tumour samples and its distribution exhibited a wide range of values. GAPDH mRNA was over-expressed in the poorly differentiated BT-20 cell line and the treatment of these cells with 1,25-dihydroxyvitamin D3 (1,25-(OH)