Searchable abstracts of presentations at key conferences in endocrinology
Endocrine Abstracts (2008) 15 P176

University of Birmingham, Birmingham, West Midlands, UK.

The pituitary tumor transforming gene binding factor (PBF) is a poorly characterised gene that is over-expressed in pituitary and thyroid tumours. Recently, we showed that subcutaneous expression of PBF elicits tumours in nude mice, and expression correlates with thyroid tumour recurrence in man. Given the established role of ionising radiation in thyroid tumourigenesis, we have now investigated the relationship between PBF and the tumour suppressor gene p53, a central component of the DNA damage checkpoint.

Glutathione S-transferase pull-down assays showed direct binding of PBF to p53. Through mutational analysis, two regions of p53 were necessary for binding PBF, between amino acids 160–318, and 318–393. Conversely, by disrupting PBF sequences, we observed that the N-terminal region of PBF is essential for p53 interaction, with binding abrogated by removal of two N-terminal regions between amino acids 28–93 and 93–149. We next showed that PBF can influence p53-mediated gene regulation through Hdm2 promoter assays in p53-null H1299 cells. PBF failed to modulate promoter activity, compared with vector-only controls, whereas p53 elicited a 143±17-fold stimulation of promoter activity. However, co-transfection with PBF significantly repressed p53 transcriptional activity (41±5-fold; P<0.001).

Exposure of p53 positive HCT116 cells to gamma-irradiation or DNA damaging agents such as etoposide produced a significant increase in PBF expression. PBF protein increased as early as 4 h after irradiation and reached maximum levels after 24 h, similar to the observed increase in p53 protein. Altogether these results are indicative of an important role for PBF in the DNA damage response through modulation of p53 activity. Furthermore, in thyroid tumours where PBF is over-expressed, we propose a novel mechanism for endocrine tumorigenesis, whereby PBF binds specifically to p53 and inhibits its regulation of critical downstream genes involved in maintaining cellular and genetic stability.

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