Sequences near the 3' secretion-specific polyadenylation site influence levels of secretion-specific and membrane-specific IgG2b mRNA in myeloma cells.
The expressed immunoglobulin gamma 2b (IgG2b) heavy-chain gene of 4T001 was cloned into the shuttle vector pSV2-gpt and transfected into myeloma J558L and lymphoma A20.2J. Northern blots indicated that the transfected gamma 2b gene was processed in a manner similar to the endogenous heavy chain in both lymphoma and myeloma cells. To identify sequences important for immunoglobulin mRNA processing, we constructed deletions around the secretion-specific polyadenylation site and introduced the deleted genes into J558L cells. The BAL deletion lacked 670 base pairs of intervening sequence between secreted and membrane regions; the Kpn deletion lacked 830 base pairs in this region. J558L cells transfected with either the entire gamma 2b gene or the delta BAL vector produced predominantly secretion-specific gamma 2b mRNA and protein. J558L cells transfected with the delta Kpn vector produced approximately equimolar amounts of secretion-specific and membrane-specific gamma 2b mRNA. Both 55,000-dalton secreted and 62,000-dalton putative surface IgG2b proteins were detected in the delta Kpn transfectants. We conclude that sequences absent in the Kpn deletion but present in the BAL deletion exert an important role in the production of secretion-specific mRNA. The Kpn deletion removes the normal site of cleavage and poly(A) addition, and it is possible that it is the absence of this site which changes the processing pattern. Alternatively, it is possible that sequences absent in the Kpn deletion but present in the BAL deletion function in regulating the production of predominantly secretion-specific mRNA in myeloma cells. The possible role of a highly conserved sequence found in this region is discussed.
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004262:DNA Restriction Enzymes; D005796:Genes; D007074:Immunoglobulin G / Q000235:genetics*; D008223:Lymphoma / Q000276:immunology; D051379:Mice; D010954:Plasmacytoma / Q000276:immunology*; D010957:Plasmids; D011061:Poly A / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism
D007074:Immunoglobulin G; D012333:RNA, Messenger; D011061:Poly A; D004262:DNA Restriction Enzymes