EXPRESSION OF CD21 AND SYNTHESIS OF ITS LIGANDS BY HELA-CELLS AFTER GROWTH IN SERUM-FREE MEDIUM

We examined the synthesis of C3 and the expression of CD21 (complement receptor type 2) by epithelial cells in serum-free culture to elimina te the effects of competing C3 fragments from serum supplementation. T he mean C3 content of Hela supernatants was twice that of lysates afte r 4 hours in culture, Immunoprecipitation of metabolically labeled pro teins from supernatants and lysates detected the structures of pro-C3 and native C3 in both supernatants and lysates, with C3 fragments iC3b and C3dg detected in supernatants only. Hybridization of total RNA wi th a human C3 complementary DNA probe revealed bands consistent with t he full-length C3 message at 5 to 6 kb, implying that the C3 fragments found in the supernatants were the result of posttranslational proces sing. Hybridization of Hela messenger RNA with complementary DNA probe s for human complement receptors revealed strong expression of CD21 me ssage but failed to detect message for CD35 (complement receptor type 1) or CD18 (beta chain of complement receptor types 3 and 4). Immunofl uorescence with the monoclonal antibodies OKB7 and HB-5 confirmed abun dant surface expression of CD21 under serum-free conditions, In a [H-3 ]thymidine uptake assay polyclonal anti-C3 inhibited Hela proliferatio n for 4 hours compared with an identical concentration of polyclonal a nti-C5 as control. Thus, in contrast to previous reports, we have obse rved abundant CD21 message at approximately 5 kb and strong surface fl uorescence with anti-CD21 monoclonal antibodies in Hela cells cultured under serum-free conditions, Serum-free conditions may help to promot e the interaction of constitutively produced C3 or C3 fragments with C D21 on malignant epithelium.