Characterization and phylogenetic epitope mapping of CD38 ADPR cyclase in the cynomolgus macaque.
BACKGROUND
The CD38 transmembrane glycoprotein is an ADP-ribosyl cyclase that moonlights as a receptor in cells of the immune system. Both functions are independently implicated in numerous areas related to human health. This study originated from an inherent interest in studying CD38 in the cynomolgus monkey (Macaca fascicularis), a species closely related to humans that also represents a cogent animal model for the biomedical analysis of CD38.


RESULTS
A cDNA was isolated from cynomolgus macaque peripheral blood leukocytes and is predicted to encode a type II membrane protein of 301 amino acids with 92% identity to human CD38. Both RT-PCR-mediated cDNA cloning and genomic DNA PCR surveying were possible with heterologous human CD38 primers, demonstrating the striking conservation of CD38 in these primates. Transfection of the cDNA coincided with: (i) surface expression of cynomolgus macaque CD38 by immunofluorescence; (ii) detection of approximately 42 and 84 kDa proteins by Western blot and (iii) the appearance of ecto-enzymatic activity. Monoclonal antibodies were raised against the cynomolgus CD38 ectodomain and were either species-specific or cross-reactive with human CD38, in which case they were directed against a common disulfide-requiring conformational epitope that was mapped to the C-terminal disulfide loop.


CONCLUSIONS
This multi-faceted characterization of CD38 from cynomolgus macaque demonstrates its high genetic and biochemical similarities with human CD38 while the immunological comparison adds new insights into the dominant epitopes of the primate CD38 ectodomain. These results open new prospects for the biomedical and pharmacological investigations of this receptor-enzyme.
Ferrero
Enza
E
Department of Genetics, Biology & Biochemistry, University of Torino, Via Santena 19 and the CeRMS Research Center for Experimental Medicine, 10126 Torino, Italy. enza.ferrero@unito.it
Orciani
Monia
M
Vacca
Paola
P
Ortolan
Erika
E
Crovella
Sergio
S
Titti
Fausto
F
Saccucci
Franca
F
Malavasi
Fabio
F
D036541:ADP-ribosyl Cyclase / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology; D051997:ADP-ribosyl Cyclase 1; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D015703:Antigens, CD / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology; D001402:B-Lymphocytes / Q000737:chemistry / Q000378:metabolism; D019556:COS Cells / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular / Q000379:methods; D003429:Cross Reactions / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics; D016678:Genome; D015894:Genome, Human; D006801:Humans; D008252:Macaca fascicularis / Q000235:genetics*; D008562:Membrane Glycoproteins; D051379:Mice; D008970:Molecular Weight; D041681:NIH 3T3 Cells / Q000737:chemistry / Q000378:metabolism; D010802:Phylogeny; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018076:DNA, Complementary; D004220:Disulfides; D000939:Epitopes; D008562:Membrane Glycoproteins; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; C498155:Cd38 protein, mouse; D051997:ADP-ribosyl Cyclase 1; D004229:Dithiothreitol