To examine this further, the relative activity of isoforms 1a, 1b and 1c on the ERBB2 promoter was also compared in the selleck chem FTY720 presence of different combinations of the CITED p300 CBP cofactors. In each case, isoform 1a was the weakest activator. TFAP2A isoform 1c expression increases in tamoxifen resistant lines and tumours Since we observed that AP 2a isoforms have differential transactivation activity on the ERBB2 promoter, we decided to investigate their expression in a biological context. ErbB2 overexpression is associated with resis tance to the oestrogen receptor antagonist tamoxi fen, since signalling from the receptor can promote oestrogen independent activation of the ER. We, therefore, investigated whether AP 2a iso form levels were altered in tamoxifen resistant lines and tumour samples.

Given that isoform 1a was the weakest activator of ERBB2 expression, we would predict that increased expression Inhibitors,Modulators,Libraries of the other isoforms might be associated with the tamoxifen resis tant phenotype. In three independent TR lines, which were cultivated in the presence of tamoxifen for at least six months, we observed a pronounced increase in levels of ErbB2 compared to the wt controls. In the same three lines, levels of iso form 1c compared to isoform 1a increased significantly, as measured by quantifying levels in Western blots as the ratio between the two isoforms. This finding led us to compare TFAP2A isoform expres sion levels in a small series of mRNA samples from TR tumours. Good quality RNA from frozen samples was required to detect isoforms levels with sufficient sensitiv ity, and this limited the number of samples available.

In these samples, ERBB2 mRNA levels tended to be higher compared to normal breast samples. AP 2a isoform 1c levels were also higher in TR tumours compared to normal breast. No significant change was observed for iso form 1a or 1b. Moreover, the difference in ratio between levels of isoform 1c and 1a in the TR tumours compared with Inhibitors,Modulators,Libraries normal breast was also significant. This would suggest a selective and differential reg ulation of AP 2a isoforms in tamoxifen resistant tumours. Discussion A significant proportion of the reports investigating the biological function of AP 2a are overexpression studies which analyse exclusively the first isoform cloned.

The existence of additional TFAP2A transcripts deriving from alternative first exons has been described in Inhibitors,Modulators,Libraries some mammals, but there has been little Inhibitors,Modulators,Libraries Inhibitors,Modulators,Libraries attempt to determine their importance or function. Our in silico analysis con sellckchem firmed that different isoforms of AP 2a occur in humans, providing at the same time evidence of a strong conservation throughout vertebrate evolution. Furthermore, ESTs for isoforms 1a and 1b can also be found for the paralogous gene AP 2b. In contrast, we were not able to identify alternative 5 variants for AP 2g, which is the homolog of AP 2a isoform 1a.