Molecular identification of a novel seven-transmembrane-domain protein, PB99, expressed at an early stage of B-cell development

Abstract

Seven-transmembrane-domain receptors (STRs) comprise the single largest family of cell surface receptors which transmit signals into cells by coupling with heterotrimeric G-proteins; (hence they are alternatively called G-protein coupled receptors, GPCR), and play versatile functions in the body. Some STRs were demonstrated to be involved in the production and/or migration of B-lymphocytes, the cells of the immune system that mediate humoral immune responses. A novel STR gene, PB99, was isolated and was found to be expressed only in cell lines representing the pre-B stage of B-lymphocyte development, which can be divided into four consecutive stages: pro-B, pre-B, mature B, and plasma cells. Full-length mouse and human PB99 cDNAs were identified and sequenced. Both mouse and human cDNAs encode STRs consisting of a relatively long N-terminal extracellular domain with a leader peptide, followed by a region containing seven transmembrane segments, and ending with a short serine/threonine rich C-terminal cytoplasmic domain. Comparison of the deduced amino, acid sequences showed that human PB99 is 78.6% homologous to mouse PB99. The high conservation during evolution suggests that PB99 performs some critical function. Furthermore, the detection of PB99 protein in bone marrow suggests that the gene is functional in the body. Comparison of PB99 to other STRs indicates that it represents a novel subgroup within a group of STRs which bind peptide ligands. The structural similarity and the sequence identity with GPCR suggest that PB99 may transmit a signal via heterotrimeric G-proteins. PB99 is a cell surface molecule with the N-terminus on the extracellular side, as shown by flow cytometric analysis of HEK293 cells transfected with HA epitope-tagged human PB99. The upregulation of PB99 expression in pre-B cells, first observed in transformed mouse pre-B cell lines, was confirmed in normal cells by quantitative RT-PCR, performed with purified cell populations representing different stages of human B-lymphocyte development. The specific increase in PB99 expression in the early stage of B-cell development suggests that this novel STR plays a signaling role during B-cell lymphopoiesis. Surprisingly, PB99 was also found to be highly expressed in non-B-lineage bone marrow cells, indicating that the gene functions in multiple hematopoietic cell lineages.