The nuclear phosphoprotein p53 is a negative regulator of cell proliferation and transformation. Loss of its normal function has been implicated in the pathogenesis of a wide array of human cancers. In thyroid tumorigenesis, p53 alterations are almost exclusively detected, at high frequency, in poorly differentiated and anaplastic carcinomas. We have recently shown that exogenous p53 re-expression in a human thyroid anaplastic carcinoma cell line (ARO), exhibiting impaired wt-p53 activity, is able to strongly inhibit cell proliferation, to abolish in vitro tumorigenicity and to induce partial restoration of thyroid differentiation properties. In particular, recovery of p53 function was able to restore ARO cell responsiveness to TSH with increased expression of the thyroid specific markers thyroglobulin (Tg), thyroperoxidase (TPO) and TSH receptor (TSH-R). With the aim of investigating the molecular mechanisms underlying this phenomenon, we evaluated the expression of the thyroid-specific transcription factors TTF-1 and PAX-8. Semi-quantitative RT-PCR analysis of mRNA isolated from ARO cells and p53-transfected ARO clones (ARO-tsp53), cultured either in the absence or presence of TSH (10mU/ml), was performed. No expression of the thyroid transcription factors was observed, both in ARO parental and ARO-tsp53 clones. These findings indicate that re-expression of Tg, TPO and TSH-R in ARO-tsp53 clones, following TSH treatment, is not mediated by modulation of their specific transcription activators. Since induction of the TSH transduction pathway results in increased intracellular cAMP levels which, in turn, leads to increased phosphorilation of CREB protein, we analyzed CREB activity in different ARO cell clones. Transient transfection assays were performed by using plasmid vectors containing a CAT reporter gene driven by a CREB-responsive promoter. A strong reduction of CREB activity was detected in ARO-tsp53 clones as compared to controls. Following TSH stimulation, re-induction of CREB activity was observed only in ARO-tsp53 clones, suggesting that recovery of p53 function is able to "normalize" intracellular signalling pathways, thus rendering anaplastic thyroid carcinoma cells again responsive to thyroid physiological stimulus. Exploitation of these p53 effects in vivo may envisage new therapeutic strategies in the treatment of thyroid undifferentiated carcinomas.
Recovery of TSH responsiveness in human thyroid anaplastic carcinoma cells following wild type p53 re-expression
1997-01-01
Abstract
The nuclear phosphoprotein p53 is a negative regulator of cell proliferation and transformation. Loss of its normal function has been implicated in the pathogenesis of a wide array of human cancers. In thyroid tumorigenesis, p53 alterations are almost exclusively detected, at high frequency, in poorly differentiated and anaplastic carcinomas. We have recently shown that exogenous p53 re-expression in a human thyroid anaplastic carcinoma cell line (ARO), exhibiting impaired wt-p53 activity, is able to strongly inhibit cell proliferation, to abolish in vitro tumorigenicity and to induce partial restoration of thyroid differentiation properties. In particular, recovery of p53 function was able to restore ARO cell responsiveness to TSH with increased expression of the thyroid specific markers thyroglobulin (Tg), thyroperoxidase (TPO) and TSH receptor (TSH-R). With the aim of investigating the molecular mechanisms underlying this phenomenon, we evaluated the expression of the thyroid-specific transcription factors TTF-1 and PAX-8. Semi-quantitative RT-PCR analysis of mRNA isolated from ARO cells and p53-transfected ARO clones (ARO-tsp53), cultured either in the absence or presence of TSH (10mU/ml), was performed. No expression of the thyroid transcription factors was observed, both in ARO parental and ARO-tsp53 clones. These findings indicate that re-expression of Tg, TPO and TSH-R in ARO-tsp53 clones, following TSH treatment, is not mediated by modulation of their specific transcription activators. Since induction of the TSH transduction pathway results in increased intracellular cAMP levels which, in turn, leads to increased phosphorilation of CREB protein, we analyzed CREB activity in different ARO cell clones. Transient transfection assays were performed by using plasmid vectors containing a CAT reporter gene driven by a CREB-responsive promoter. A strong reduction of CREB activity was detected in ARO-tsp53 clones as compared to controls. Following TSH stimulation, re-induction of CREB activity was observed only in ARO-tsp53 clones, suggesting that recovery of p53 function is able to "normalize" intracellular signalling pathways, thus rendering anaplastic thyroid carcinoma cells again responsive to thyroid physiological stimulus. Exploitation of these p53 effects in vivo may envisage new therapeutic strategies in the treatment of thyroid undifferentiated carcinomas.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
