Role of the second extracellular loop of human C3a receptor in agonist binding and receptor function

Producción CientíficaThe C3a anaphylatoxin receptor (C3aR) is a G protein-coupled receptor with an unusually large second extra-cellular loop (e2 loop,;172 amino acids). To determinethe function of this unique structure, chimeric and de-letion mutants were prepared and analyzed in trans-fected RBL-2H3 cells. Whereas replacement of the C3aRN-terminal segment with that from the human C5a re-ceptor had minimal effect on C3a binding, substitutionof the e2 loop with a smaller e2 loop from the C5a recep-tor (C5aR) abolished binding of125I-C3a and C3a-stimu-lated calcium mobilization. However, as much as 65% ofthe e2 loop sequence (amino acids 198 –308) may be re-moved without affecting C3a binding or calcium re-sponses. The e2 loop sequences adjacent to the trans-membrane domains contain multiple aspartate residuesand are found to play an important role in C3a bindingbased on deletion mutagenesis. Replacement of five as-partate residues in the e2 loop with lysyl residues sig-nificantly compromised both the binding and functionalcapabilities of the C3a receptor mediated by intact C3aor by two C3a analog peptides. These data suggest atwo-site C3a-C3aR interaction model similar to that es-tablished for C5a/C5aR. The anionic residues near the Nand C termini of the C3aR e2 loop constitute a non-effector secondary interaction site with cationic resi-dues in the C-terminal helical region of C3a, whereas theC3a C-terminal sequence LGLAR engages the primaryeffector site in C3aR

The rate of protein synthesis in hematopoietic stem cells is limited partly by 4E-BPs

Adult stem cells must limit their rate of protein synthesis, but the underlying mechanisms remain largely unexplored. Differences in protein synthesis among hematopoietic stem cells (HSCs) and progenitor cells did not correlate with differences in proteasome activity, total RNA content, mRNA content, or cell division rate. However, adult HSCs had more hypophosphorylated eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP2 as compared with most other hematopoietic progenitors. Deficiency for 4E-BP1 and 4E-BP2 significantly increased global protein synthesis in HSCs, but not in other hematopoietic progenitors, and impaired their reconstituting activity, identifying a mechanism that promotes HSC maintenance by attenuating protein synthesis.National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant R01 DK100848)Cancer Prevention and Research Institute of Texa

CCR5 Blockade Modulates Inflammation and Alloimmunity in Primates

Pharmacologic antagonism of CCR5, a chemokine receptor expressed on macrophages and activated T cells, is an effective antiviral therapy in patients with macrophage-tropic HIV infection, but its efficacy in modulating inflammation and immunity is only just beginning to be investigated. In this regard, the recruitment of CCR5-bearing cells into clinical allografts is a hallmark of acute rejection and may anticipate chronic rejection, whereas conventionally immunosuppressed renal transplant patients homozygous for a nonfunctional Δ32 CCR5 receptor rarely exhibit late graft loss. Therefore, we explored the effects of a potent, highly selective CCR5 antagonist, Merck\u27s compound 167 (CMPD 167), in an established cynomolgus monkey cardiac allograft model. Although perioperative stress responses (fever, diminished activity) and the recruitment of CCR5-bearing leukocytes into the graft were markedly attenuated, anti-CCR5 monotherapy only marginally prolonged allograft survival. In contrast, relative to cyclosporine A monotherapy, CMPD 167 with cyclosporine A delayed alloantibody production, suppressed cardiac allograft vasculopathy, and tended to further prolong graft survival. CCR5 therefore represents an attractive therapeutic target for attenuating postsurgical stress responses and favorably modulating pathogenic alloimmunity in primates, including man

CCR5 blockade modulates inflammation and alloimmunity in primates

Pharmacologic antagonism of CCR5, a chemokine receptor expressed on macrophages and activated T cells, is an effective antiviral therapy in patients with macrophage-tropic HIV infection, but its efficacy in modulating inflammation and immunity is only just beginning to be investigated. In this regard, the recruitment of CCR5-bearing cells into clinical allografts is a hallmark of acute rejection and may anticipate chronic rejection, whereas conventionally immunosuppressed renal transplant patients homozygous for a nonfunctional Δ32 CCR5 receptor rarely exhibit late graft loss. Therefore, we explored the effects of a potent, highly selective CCR5 antagonist, Merck\u27s compound 167 (CMPD 167), in an established cynomolgus monkey cardiac allograft model. Although perioperative stress responses (fever, diminished activity) and the recruitment of CCR5-bearing leukocytes into the graft were markedly attenuated, anti-CCR5 monotherapy only marginally prolonged allograft survival. In contrast, relative to cyclosporine A monotherapy, CMPD 167 with cyclosporine A delayed alloantibody production, suppressed cardiac allograft vasculopathy, and tended to further prolong graft survival. CCR5 therefore represents an attractive therapeutic target for attenuating postsurgical stress responses and favorably modulating pathogenic alloimmunity in primates, including man