MODIFIED BINDING OF PROTEINS FROM CALCITONIN-NEGATIVE TUMOR-CELLS TO THE NEUROENDOCRINE-SPECIFIC CANNTG MOTIF OF THE CALCITONIN-GENE

Transcription of the calcitonin (CT) gene in the medullary thyroid car cinoma (MTC) cell line TT is modulated by a neuroendocrine-specific en hancer fragment (nucleotides - 965 to - 905) containing two CANNTG mot ifs (E2 and E3) and an Ets-like response element. To determine the cel l-specific component of this fragment, oligonucleotides containing the individual elements were inserted in front of a minimal CT promoter a nd tested for reporter protein production in CT-positive (TT) and -neg ative (RO-D81 and HeLa) cells. In TT cells, using two copies of E2 or four copies of Ets increased minimal promoter activity a 20-40 fold. U sing two copies of E3 had no effect on minimal promoter activity. In C T-negative MTC cells (RO-D81), the Ets response element was active but the two copies of E2 were not. Similar results were obtained with the non-neuroendocrine cell-line HeLa. I therefore concluded that E2 was the cell-type-specific component of the enhancer. An E2-specific bindi ng protein was detected in both MTC cell lines but not in HeLa. This p rotein had different mobility and DNA-binding specificity in CT-positi ve TT cells and CT-negative RO-D81 cells. In conclusion, the CAGCTG mo tif of E2 modulated the cell-specific transcription of the CT gene, an d its inactivation in CT-negative MTC cells correlated with modificati ons in its binding proteins.