THE EXTENDED HINGE REGION OF IGG3 IS NOT REQUIRED FOR HIGH PHAGOCYTICCAPACITY MEDIATED BY FC-GAMMA-RECEPTORS, BUT THE HEAVY-CHAINS MUST BEDISULFIDE-BONDED

Fcgamma receptor (FcgammaR) phagocytosis and respiratory burst were in duced by chimeric mouse-human anti-(4-hydroxy-5-iodo-3-nitrophenyl) ac etyl IgG3 antibodies with mutations in hinge and/or in C(H)1 region. I gG3 mutants with different hinge length ranging from 47 to 0 amino aci ds, an IgG3 molecule with an artificial hinge of just one cysteine res idue (HM-1), and two hybrid IgG3 molecules with IgG4 hinge or IgG4 C(H )1-hinge were tested. Using the monocytic cell line U937 as effector c ells, the mutated IgG3 molecules were very similar, revealing high act ivity, while the IgG3/IgG4 hybrids revealed a slightly reduced activit y. However, the hingeless (0-h) mutant was negative, except after inte rferon-gamma stimulation when it became slightly positive. Interesting ly, HM-1 was as active as the IgG3 mutants. With polymorphonuclear leu cocytes (PMN) as effector cells we obtained some day-to-day variations , but all the IgG3 mutants were highly active, with the two shortest h inge mutants somewhat less active. The IgG3/IgG4 hybrid molecules reve aled an intermediate activity, while IgG4 wild-type and the 0-h mutant were negative. However, the HM-1 molecule revealed an activity simila r to that of the IgG3 mutants. The phagocytic activity of U937 was inh ibited by monomeric IgG, indicating the importance of FcgammaRI. In co ntrast, with PMN both blockage of FcgammaRII and cleavage of FcgammaRI II were required to significantly reduce the phagocytosis and respirat ory burst, thus showing that both receptors contribute to the effect. These results demonstrate that the extended IgG3 hinge region is not n ecessary for a high phagocytic activity and that the major structural importance of the hinge is to connect the two heavy chains in this reg ion.