In vitro study of IgM polymerization

The polymerization of 7S IgMs from normal rabbit lymphoid cells, stimulated either with antigen or with mitogen (Con A), has been studied. The process was analyzed by characterizing the various molecular forms by sucrose gradient sedimentation and susceptibility to anti-μ serum and 2-mercaptoethanol. It has been shown that native J chain and an enzyme are both required for the proper assembly of IgM pentamer. The enzyme preparation (PMF) is active only if it is extracted from spleen cells stimulated to IgM production. When the extract is prepared from nonstimulated lymphoid cells, or from liver cells, incubation of IgMs with PMF does not lead to the formation of 19S IgM, but to molecules of intermediate size and to various aggregates. It is shown that antibody activity of IgMs and of these heterogeneous polymers are not susceptible to treatment with 2-mercaptoethanol. In contrast, antibody activity of the pentameric IgM is completely inhibited by 2-mercaptoethanol. A PMF inhibitory substance was present in the postmicrosomal supernatant. When added in the incubation medium, this substance prevented the proper polymerization. Its eventual role in IgM biosynthesis in nonstimulated, and specifically stimulated cells is discussed compared with mitogen stimulated cells, and tumor lymphoid cells.