CXCR7 influences the migration of B cells during maturation.

The atypical chemokine receptor CXCR7 binds the chemokines CXCL12 and CXCL11. The receptor is widely expressed and was shown to tune CXCR12-induced responses of CXCR4. Here, the function of CXCR7 was examined at late stages of human B-cell maturation, when B cells differentiate into Ab-secreting plasmablasts. We identified two populations of CXCR7(+) cells in tonsillar lymphocytes, one being presumably memory B cells or early plasmablasts (FSC(low) CD19(+) CD38(mid) ) and the other being plasmablasts or early plasma cells (FSC(high) CD19(+) CD38(+) ). CXCR7 is expressed on CD19(+) CD27(+) memory B cells, on CD19(+) CD38(+) CD138(-) and intracellular immunoglobulin high plasmablasts, but not on CD19(+) CD138(+) icIg(high) plasma cells. The differential expression pattern suggests a potential contribution of the scavenger receptor in final B-cell maturation. On in vitro differentiating B cells, we found a marked inverse correlation between CXCR7 and CXCR5 cell surface levels, whereas expression of CXCR4 remained almost constant. Migration assays performed with tonsillar mononuclear cells or in vitro differentiated cells revealed that inhibition of CXCR7 markedly increases chemotaxis toward CXCL12, especially at late stages of B-cell maturation. Chemotaxis was attenuated in the presence of CXCR4 antagonists, confirming that migration is CXCR4 mediated. Our findings unequivocally demonstrate a novel role for CXCR7 in regulating the migration of plasmablasts during B-cell maturation.

Complementary methods provide evidence for the expression of CXCR7 on human B cells.

PTMs of extracellular domains of membrane proteins can influence antibody binding and give rise to ambivalent results. Best proof of protein expression is the use of complementary methods to provide unequivocal evidence. CXCR7, a member of the atypical chemokine receptor family, mainly functions as scavenger for the chemokines CXCL12 and CXCL11. The expression of CXCR7 on nonhematopoietic cells and neoplasms is widely accepted, however, its expression on leukocytes was recently challenged. To solve the dissent, we thoroughly analyzed the expression of CXCR7 on human B cells. We validated the efficiency of different epitope-specific monoclonal antibodies to detect CXCR7 on transfected cells and primary human B cells. The specificity of the used antibodies was further confirmed by an experimentally independent double labeling approach. Examination of CXCR7-dependent scavenging of fluorescent-labeled CXCL12 revealed functional expression of the receptor on human B cells. Moreover, real-time PCR analysis of CXCR7 mRNA showed the presence of transcripts in human leukocytes. Finally, two CXCR7-specific peptides were identified by MS in immunoprecipitates from primary human B cells. Thus, we present a strategy based on combined proteomic and functional approaches that can be used to solve dissents on protein expression, i.e. demonstrating the expression of CXCR7 on human leukocytes.

Interactions in the pollen-specific receptor-like kinases-containing signaling network.

The pollen-specific receptor-like kinases (PRKs) from Solanum lycopersicum, LePRK1 and LePRK2, are believed to be involved in the regulation of pollen germination and pollen tube growth. They appear to be part of a multimeric complex in which the transmembranic LePRKs presumably have a key position in transducing exogenous signals through the plasma membrane. Here, we focused on extra- and intracellular interactions involving the LePRKs. We show in yeast two-hybrid experiments a cross-interaction of putative PRK-ligands, the oligomerization of LePRK2 and a direct contact of LePRKs to activated Rho proteins of plants (ROPs). Moreover, we observed that pollen-specific RopGEFs, which catalyze ROP activation and may be regulated by PRK interaction, are active in vitro while autoinhibition seems to occur in vivo. We suggest that activation of RopGEFs as a checkpoint in PRK signal transduction is a more complex event including further components in planta. Our findings point to some new aspects in PRK-mediated signal transduction implying a LePRK2 complex with different signaling activity and a further direct control of LePRKs by activated ROP.