Immunoglobulin heavy chain mRNA in mitogen-stimulated B cells.
This paper relates the synthesis of DNA, immunoglobulin and heavy chain (H) mRNA in murine spleen cells following activation of B cells with lipopolysaccharide from E. coli (LPS). Spleen cells (CBA/H mice) were cultivated with 10% FCS and 10 mug LPS/ml. 4 h pulses with [3H]thymidine showed that DNA synthesis was stimulated within the first day following LPS activation and exhibited a sharp peak at 24 h. The shape of the DNA synthesis curve suggests that the cells susceptible to LPS stimulation are activated in a synchronous manner. Stimulation of H-chain mRNA (H-mRNA) synthesis proceeded rapidly (within 6 h of LPS addition) and peaked around 24 h, in parallel to DNA synthesis. The H-mRNA was isolated and quantitated by making use of its interaction with IgG[1, 2]. The actual level of H-mRNA in the culture increased threefold during the first 24 h and then doubled within the next 48 h. Estimates of the actual number of H-mRNA were approximately 200 molecules H-m-RNA/cell on day 0 rising to 1800/cell on day 3. In such a mixed cell population these figures will be accurate only within a factor of 2-3 (at least 35% B cells in spleen cell suspensions at the commencement of the culture, with up to 35-60% of plasma blasts by day 3 and 4 of LPS treatment). Translation of the lymphoid cell mRNA in oocytes from Xenopus laevis demonstrated that stimulation of H-mRNA synthesis was restricted to mu-mRNA, although some gamma-mRNA was present in the original spleen cells. High levels of synthesis of immunoglobulin followed after a lag period of about 24 h following LPS addition peaking after 48 and 72 h; the proportional Ig production relative to total protein synthesis reached 26% on days 3 and 4. Stimulation of Ig production was limited to IgM. Rapid stimulation of mitosis and H-mRNA synthesis thus precedes the maximum synthesis of Ig molecules, suggesting a translational block on H-mRNA during cell maturation. There was no apparent block on the transport of H-mRNA from the nucleus during early stages of activation.
D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis*; D005260:Female; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis*; D007148:Immunoglobulin mu-Chains / Q000096:biosynthesis; D007700:Kinetics; D008070:Lipopolysaccharides / Q000276:immunology; D008213:Lymphocyte Activation; D051379:Mice; D009865:Oocytes / Q000378:metabolism; D011135:Polysaccharides, Bacterial / Q000276:immunology; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis*; D013154:Spleen / Q000276:immunology; D014981:Xenopus
D007143:Immunoglobulin Heavy Chains; D007148:Immunoglobulin mu-Chains; D008070:Lipopolysaccharides; D011135:Polysaccharides, Bacterial; D012333:RNA, Messenger; D004247:DNA