2 MURINE HOMOLOGS OF THE HUMAN CHEMOKINE RECEPTOR CXCR4 MEDIATING STROMAL CELL-DERIVED FACTOR 1-ALPHA ACTIVATION OF G(12) ARE DIFFERENTIALLY EXPRESSED IN-VIVO

Previous results have shown that pertussis toxin-sensitive G(i) protei ns are likely to be involved in regulating the emigration of mature th ymocytes from the thymus. In this study, a low stringency polymerase c hain reaction (PCR) approach was used to identify G(i) protein-coupled cell surface receptors expressed in mouse thymocytes. Among the ten G protein-coupled receptor cDNA isolated, the most prevalent cDNA encod ed a polypeptide highly homologous to the human leukocyte-expressed se ven-transmembrane-domain receptor LESTR, also referred to as HIV entry cofactor, fusin, or CXCR4. Isolation of full-length cDNA revealed tha t alternative RNA splicing produces transcripts encoding two isoforms of the murine LESTR, differing by the presence of two amino acids in t he N-terminal portion of the longer protein. Functional reconstitution of recombinant murine LESTR with recombinant heterotrimeric G protein s in baculovirus-infected insect cells showed that both receptor varia nts mediate stromal cell-derived factor 1 alpha activation of the pert ussis toxin-sensitive G protein G(i2). Receptor subtype-specific rever se transcriptase-PCR analysis revealed differential expression of the two receptor mRNA in lymphoid tissues and brain, indicating that disti nct functions are mediated by the two receptor isoforms in these tissu es. The presence of LESTR mRNA in very early thymocytes as well as in immature (CD4(+) CD8(+)) thymocytes suggests that both CD4 and LESTR a re co-expressed and render developing human thymocytes susceptible for HIV entry which may affect generation of both CD4(+) CD8(-) and CD4(- ) CD8(+) mature lineages.