TISSUE-SPECIFIC PROMOTERS OF THE ALPHA-HUMAN FOLATE RECEPTOR GENE YIELD TRANSCRIPTS WITH DIVERGENT 5'-LEADER SEQUENCES AND DIFFERENT TRANSLATIONAL EFFICIENCIES

The ct human folate receptor (alpha hFR), or KB cell folate receptor, gene contains two major promoters that produce transcripts, KB1 and KB 4, varying only in the length and sequence of their 5' untranslated re gions (UTRs). Using RNase protection assays specific for each isoform, we show that the level of expression of these two transcripts is tiss ue-specific, indicating that promoter usage is regulated, not constitu tive. RNA stabilities and translational efficiencies of the KB1 and KB 4 transcripts were compared to determine the functional significance o f the different 5' UTRs. Analyses of RNA turnover in vivo with actinom ycin D to block new transcription and in vitro with a cytoplasmic extr act indicate no discernible differences in the stabilities of the two transcripts. However, the KB4 transcript is 2-3-fold mon efficiently t ranslated in wheat germ extracts in vitro and transfected CHO cells in vivo. Also, high ionic strength, which favours the formation of RNA s econdary structure, differentially affects the translational efficienc ies of the two transcripts. Translation of the longer KB1 mRNA is 2-5- fold more inhibited by hypertonic conditions than translation of the K B4 mRNA. Because the 5' UTR of KB1 is approximately four times longer than the 5' UTR of KB4, 149 bp (75%) of the KB1 5' UTR were deleted to determine whether the long leader sequence inhibited translation. The resulting derivative, dKB1, has a 5' UTR similar in length, but not s equence, to the 5' UTR of KB4. dKB1 is translated at a level approachi ng that of KB4 in wheat germ extracts, indicating that the upstream po rtion of the 5' leader sequence contributes to the relative translatio nal inefficiency of KB1. Hence, one consequence of tissue-specific pro moter usage is the production of alpha hFR transcripts with different 5' non-coding regions that affect translational efficiency.