Human FcγRIII (CD16), a low-affinity receptor expressed on several different cell types, has a polymorphism on polymorphonuclear cells (FcγRIII(PMN)) identified by the NA1 and NA2 markers. Inasmuch as this polymorphism has functional consequences, an understanding of the structural biology of FcγRIII may provide important insight into receptor function. We analyzed FcγRIII(PMN) by SDS-PAGE and found that receptor from individuals allotyped for either NA1 or NA2 contained only one protein after removal of N-linked glycosylations (19 and 21 kDa respectively) whereas receptor from NA1/2 individuals contained both bands. Because some reports indicate that digestion of FcγRIII on NK cells (FcγRIII(NK)) with N-glycanase also results in two bands on SDS-PAGE, we investigated FcγRIII(NK) to explore the possibility of a corresponding allelic polymorphism in this receptor. Contrary to expectation FcγRIII(NK) from all donors irrespective of their NA allotype contained two bands (20 and 24 kDa) on SDS-PAGE after deglycosylation. In addition, those distinct epitopes on the extracellular domain of FcγRIII(PMN) found with mAb B73.1 and CLB gran 11 in association with the NA allotypic differences are expressed (or not expressed) on FcγRIII(NK) independent of donor NA allotype. FcγRIII(PMN) and FcγRIII(NK) differ at the protein level as they have different m.w. (glycosylated and deglycosylated), different epitopes in the extracellular domain (not attributable to tissue-specific glycosylation), and differential expression of the NA allelic protein polymorphism. Although the membrane anchor of FcγRIII(PMN) is phosphatidylinositol-specific phospholipase C sensitive glycosyl-phosphatidylinositol linkage, FcγRIII(NK) is insensitive to phosphatidylinositol-specific phospholipase C. However, a form of FcγRIII(NK) is released from NK cells upon incubation at 37°C. Thus, the basis for the two bands in FcγRIII(NK) after N-linked deglycosylation is neither coexpression of two molecular isoforms with different membrane anchors nor an identifiable allelic polymorphism in m.w. restricted to FcγRIII(NK) (p < 10-6). The differences between the two receptors indicate that, independent of cell anchor type, PMN and mononuclear cells must have different molecular isoforms. The allelic variants, different isoforms, alternative anchor mechanisms and release processes provide for an extensive genetic and regulatory diversity in FcγRIII function.