MOLECULAR-CLONING AND MOLECULAR BIOLOGICAL STUDIES OF HUMAN THYMIDYLATE SYNTHASE GENE

Thymidylate synthase (TS) provides the only de novo source of thymidylate for deoxyribonucleic acid (DNA) synthesis and is a key target of cancer chemotherapeutic agents. Expression of its activity is strongly dependent on the cell cycle. Therefore, it is important for understanding the abnormal growth of cancer cells as well as for cancer chemotherapy to elucidate the structure of a TS gene and the regulatory mechanism of expression of its gene at the molecular level. From this point of view, genomic DNA segments partially coding for human TS were cloned from the mouse cell transformant obtained by gene transfer. By using them as a probe, functional cDNA and genomic DNA (approx. 18 kb) clones for human TS were isolated, and the entire nucleotide sequence of these cloned DNAs including all the introns determined. On the basis of the sequence data, we revealed an amino acid sequence of human TS, and organization and several structural features of its gene were revealed. Then, multiple transcription initiation sites and transcriptional regulatory elements of the TS gene were identified. Furthermore, the importance of the post-transcriptional regulation in the cell-cycle dependent expression of the gene was shown. The results obtained so far have opened the way to elucidate the molecular mechanism regulating the TS gene expression entirely.