title: Tracking membrane and secretory immunoglobulin alpha heavy chain mRNA variation during B-cell differentiation by real-time quantitative polymerase chain reaction.
abstract: Primary transcripts for all Ig heavy chain isotypes are alternatively processed to encode either secreted or membrane forms of the same antibody and, in plasma cells, a shift towards the secreted form occurs. In principle, measuring the relative quantities of secreted and membrane forms for a particular isotype could monitor B-cell plasmacytoid differentiation. Ratios of alpha heavy chain mRNA secreted (alphas) to membrane (alpham) form were assessed by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR; TaqMan) using an IgA plasma cell line (NCI-H929), a surface IgA+ line (Dakiki) and human tonsillar B cells. While NCI-H929 cells showed the highest alphas: alpham ratio as expected, alphas mRNA predominated for all unstimulated B cells and Dakiki cells. Treatment of B cells and Dakiki cells with IL-2 and IL-10 resulted in a further progression towards the alphas form, correlating with increased human plasma cell antigen-1 (HPC1) mRNA levels. However, alpha mRNA processing and HPC1 expression were independently regulated, as IFN-gamma treatment suppressed HPC1 levels while increasing alphas: alpham ratios. Cytokine-mediated increases in the alphas: alpham ratio resulted from strongly enhanced levels of alphas with relatively constant alpham values. Differentiation-related changes in mRNA processing can thus be tracked by automated quantitative PCR.
authors.affiliation: The Institute for Immunology and Allergy Research, Westmead Hospital, New South Wales, Australia.
authors.forename: S D
authors.initials: SD
authors.lastname: Xiang
authors.forename: E M
authors.initials: EM
authors.lastname: Benson
authors.forename: I S
authors.initials: IS
authors.lastname: Dunn
mesh_terms: D000944:Antigens, Differentiation, B-Lymphocyte / Q000235:genetics / Q000378:metabolism; D001402:B-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D002454:Cell Differentiation; D002469:Cell Separation; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D007070:Immunoglobulin A / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007071:Immunoglobulin A, Secretory / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology; D016753:Interleukin-10 / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
chemical_list: D000944:Antigens, Differentiation, B-Lymphocyte; D007070:Immunoglobulin A; D007071:Immunoglobulin A, Secretory; D007143:Immunoglobulin Heavy Chains; D007376:Interleukin-2; D012333:RNA, Messenger; D016753:Interleukin-10; D007371:Interferon-gamma