FOLATE RECEPTOR-TYPE-GAMMA IS PRIMARILY A SECRETORY PROTEIN DUE TO LACK OF AN EFFICIENT SIGNAL FOR GLYCOSYLPHOSPHATIDYLINOSITOL MODIFICATION - PROTEIN CHARACTERIZATION AND CELL-TYPE SPECIFICITY

A novel isoform of the human folate receptor (FR, type gamma) was rece ntly identified in hematopoietic tissues [Shen et al. (1994) Biochemis try 33, 1209-1215]. In that report, Cos-1 cells, transiently transfect ed with the cDNA for FR-gamma, produced relatively poor expression of the receptor on the cell surface. In this study, several recombinant C hinese hamster ovary (CHO) cell lines were produced by stable transfec tion with the cDNA for FR-gamma followed by amplification. Similar rec ombinant CHO cell lines were produced that expressed the glycosylphosp hatidylinositol- (GPI-) anchored FR type beta and a truncated form of FR type beta (FR-beta Delta), in which the normal carboxyl-terminal si gnal for GPI anchor attachment was deleted. Both FR-gamma- and FR-beta Delta-expressing CHO cells produced a [H-3]folic acid binding protein in the medium with a similar time course over a 24-h period; in contr ast to intact FR-beta, relatively insignificant amounts of either FR-g amma or FR-beta Delta were associated with the CHO cell surface and th is was unaltered by the absence of serum in the medium. The FR-gamma- and FR-beta Delta-producing CHO cells did not differ significantly in intracellular FR levels. Furthermore, the mRNA level for FR-gamma did not exceed that for FR-beta Delta. When deglycosylated with hydrogen f luoride, both FR-gamma and FR-beta Delta showed similar apparent molec ular weights on Western blots as predicted for the intact polypeptides . Chimeric constructs of FR-gamma and FR-beta cDNAs resulting in inter changing of the unconserved, carboxyl-terminal segments of the two pro teins were expressed in human 293 fibroblasts; the results indicate th at FR-gamma is primarily a secretory protein due to a divergence of it s carboxyl-terminal amino acid sequence from the other members of the FR gene family, resulting in an inefficient signal for GPI modificatio n. The relative affinities of the secreted FR-gamma for folic acid and the diastereoisomers of reduced folate compounds were in the order fo lic acid (K-D = 0.42 x 10(-)9 M) > (6S)/(6R)-N-5-methyltetrahydrofolat e > (6S)/(6R)-N-5-formyltetrahydrofolate. Evidence is presented that e xpression of the secreted full-length FR-gamma is restricted to certai n hematopoietic cell types. FR-gamma is, therefore, a potential serum marker for certain malignancies of hematopoietic tissues and also a ca ndidate protein for the high-affinity serum folate binder that is elev ated during folate deficiency.