Immunoglobulin μ Heavy Chains Do Not Mediate Tyrosine Phosphorylation of Igα from the ER- cis- Golgi

Abstract

Signals delivered by Ig receptors guide the development of functional B lymphocytes. For example, clonal expansion of early μ heavy chain (μHC)-positive pre-B cells requires the assembly of a signal-competent pre-B cell receptor complex (pre-BCR) consisting of a μHC, a surrogate L chain, and the signal dimer Igαβ. However, only a small fraction of the pre-BCR is transported to the cell surface, suggesting that pre-BCR signaling initiates already from an intracellular compartment, e.g., the endoplasmic reticulum (ER). The finding that differentiation of pre-B cells and allelic exclusion at the IgH locus take place in surrogate L chain-deficient mice further supports the presence of a μHC-mediated intracellular signal pathway. To determine whether a signal-competent Ig complex can already be assembled in the ER, we analyzed the consequence of pervanadate on tyrosine phosphorylation of Igα in J558L plasmacytoma and 38B9 pre-B cells transfected with either a transport-competent IgL chain-pairing or an ER-retained nonpairing μHC. Flow cytometry, combined Western blot-immunoprecipitation-kinase assays, and confocal microscopy revealed that both the nonpairing and pairing μHC assembled with the Igαβ dimer; however, in contrast to a pairing μHC, the nonpairing μHC was retained in the ER-cis-Golgi compartment, and neither colocalized with the src kinase lyn nor induced tyrosine phosphorylation of Igα after pervanadate treatment of cells. On the basis of these findings, we propose that a signal-competent Ig complex consisting of μHC, Igαβ, and associated kinases is assembled in a post-ER compartment, thereby supporting the idea that a pre-BCR must be transported to the cell surface to initiate pre-BCR signaling.