UPSTREAM ORGANIZATION OF AND MULTIPLE TRANSCRIPTS FROM THE HUMAN FOLYLPOLY-GAMMA-GLUTAMATE SYNTHETASE GENE

Folylpoly-gamma-glutamate synthetase (FPGS) is essential for the survi val of proliferating mammalian cells and central to the action of all ''classical'' folate antimetabolites. We report the isolation of cDNAs corresponding to the 5' ends of FPGS mRNA from both human and hamster cells which include a start codon upstream of and in-frame with the A UG in the previously reported FPGS open reading frame. The predicted h amster and human amino-terminal extension peptides have features consi stent with a mitochondrial targeting sequence. Ribo-nuclease protectio n and 5'-rapid amplification of cDNA ends assays indicated multiple tr anscriptional start sites consistent with the sequence of the promoter region of this gene, which was highly GC-rich and did not contain TAT A or CCAAT elements. These start sites would generate two classes of t ranscripts, one including the upstream AUG and one in which only the d ownstream AUG would be available for translation initiation. Transfect ion of the full length human cDNA into cells lacking FPGS restored the ir ability to grow in the absence of glycine, a product of mitochondri al folate metabolism, as well as of thymidine and purines. Therefore, we propose that the mitochondrial and cytosolic forms of FPGS are deri ved from the same gene, arising from the use of the two different tran slation initiation codons, and that the translation products differ by the presence of a 42-residue amino-terminal mitochondrial leader pept ide.