Role for p300 in Pax 8 Induction of Thyroperoxidase Gene Expression

The nuclear p300 protein functions as a co-activator of gene transcription. Here we show that p300 works as a co-activator of the transcription factor Pax 8 on the thyroperoxidase gene promoter. Consistent with its role as co-activator, p300 potentiates Pax 8-activated transcription. Furthermore, we provide evidence supporting the formation of a complex between both factors in vivo and in vitro. This interaction involves the amino-terminal and CH3 domains of p300 and the trans-activation domain of Pax 8 at its carboxyl-terminal end. We show that the CH3 domain is crucial for the co-activator role of p300 on the thyroperoxidase gene promoter. In agreement with our finding and with the ability of the adenoviral protein E1A to bind p300, we show that E1A down-regulates Pax 8 activity

Identification of the minimal melanocyte-specific promoter in the melanocortin receptor 1 gene

Abstract Background The understanding of cutaneous pigmentation biology is relevant from the biologic and clinical point of view. The binding of α-melanocortin and its specific receptor, on the plasma membrane of melanin synthesising cells, plays a crucial role in melanins biosynthesis. Furthermore, loss of MC1R function is associated with an increased incidence of melanoma and non-melanoma skin cancer. The expression of the α-melanocortin receptor gene is highly controlled but, at the present, region responsible for tissue-specific activity of the gene promoter has not been identified. Methods We have cloned the genomic sequences upstream the human MC1R coding gene. A DNA fragment of 5 kilobases upstream the human MC1R encoding sequence was placed in front of a reporter gene and several deletion mutants of such fragment have been prepared. These constructs have been tested for the ability to drive the melanocyte-specific gene expression of the reporter gene using transfection experiments in melanocyte and non-melanocyte cell lines. From these experiments we identified a DNA fragment with the ability to drive the gene transcription in a tissue-specific way and we used this small DNA fragment in DNA-protein interaction assays. Results We show that the 150 base pairs upstream the MC1R gene initiation codon are able to drive the melanocyte-specific gene transcription. Furthermore, we provide experimental evidences suggesting that on such minimal melanocyte-specific gene promoter can assemble tissue-specific complexes. Conclusion The present results strongly imply that the transcriptional regulation of the melanocyte-specific MC1R gene requires an internal promoter located in the 150 base pairs upstream the initiation codon.</p