Synthesis of the mouse complement component C4 (Ss-protein) by peritoneal macrophages: kinetics of secretion and glycosylation of the subunits.

Abstract

AbstractSs antigen was partially purified from mouse EDTA plasma, and a highly specific antiserum was prepared. Peritoneal exudate cells from Ss‐high mouse strains were labeled in primary tissue culture with [35S]methionine. Ss protein was found to be synthesized by macrophages and to be secreted into the culture medium as shown by indirect immunoprecipitations performed with this antiserum. The unreduced precipitated Ss molecule coelectrophoresed with human C4 at an apparent mol. wt. of 200000. Upon reduction, Ss protein dissociated into three subunits α, β and γ of mol. wts. 98000, 70000 and 32000. This pattern was almost identical to the one obtained for human C4. The Ss a subunit was converted by human Cls into an α subunit of approximately 85 Kdalton. This conversion is a characteristic property of C4. A single‐chain intracellular precursor for the Ss protein (pre‐Ss: 200 Kdalton) was also immunoprecipitated. In pulse‐chase experiments, it disappeared from the cytoplasmic extract within less than 10 h in a reciprocal manner to the appearance of extracellular mature Ss protein. No significant amounts of high‐mol. wt. extracellular precursors were detected. A group of four single‐chain molecules of mol. wt. between 160000 and 220 000 was immunoprecipitated from the tissue culture supernatant with anti‐Ss antiserum. These molecules could not be chased into mature Ss protein. The α, α′ and β subunits of Ss and the α chain of C3 incorporated [3H]mannose and [3H]glucosamine, the Ss α and C3 β chains did not.