Tracking membrane and secretory immunoglobulina α heavy chain mRNA variation during B-cell differentiation by real-time quantitative polymerase chain reaction

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 α heavy chain mRNA secreted (αs) to membrane (αm) 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 αs:αm ratio as expected, αs 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 αs form, correlating with increased human plasma cell antigen-1 (HPC1) mRNA levels. However,α mRNA processing and HPC1 expression were independently regulated, as IFN-γ treatment suppressed HPC1 levels while increasing αs:αm ratios. Cytokine-mediated increases in the αs:αm ratio resulted from strongly enhanced levels of αs with relatively constant αm values. Differentiation-related changes in mRNA processing can thus be tracked by automated quantitative PCR.