Biochemical characterization of PRV-1, a novel hematopoietic cell surface receptor, which is overexpressed in polycythemia rubra vera.
The cDNA for polycythemia rubra vera 1 (PRV-1), a novel hematopoietic receptor, was recently cloned by virtue of its overexpression in patients with polycythemia vera. PRV-1 is a member of the uPAR/CD59/Ly6 family of cell surface receptors, which share a common cysteine-rich domain and are tethered to the cell surface via a glycosylphosphatidylinositol (GPI) link. We have determined the intron-exon structure of the PRV1 gene and show that the locus is structurally intact in patients with polycythemia vera. Thus, PRV-1 overexpression in these patients is not due to rearrangement or structural alteration of the gene. Northern blot analysis detects multiple PRV-1 transcripts. Here we show that these transcripts arise from alternative polyadenylation and encode the same protein. Biochemical analysis reveals that PRV-1 is N-glycosylated and embedded in the cell membrane by a lipid anchor, like other members of this family. Moreover, PRV-1 is shed from the cell surface because soluble protein can be detected in cell supernatants. Fluorescence-activated cell sorting analysis of stably transfected cells revealed that PRV-1 is recognized by antibodies directed against the neutrophil antigen NB1/CD177. Flow cytometry of bone marrow and peripheral blood of both healthy donors and patients with polycythemia vera showed that PRV-1 protein is expressed on myeloid cells of the granulocytic lineage. However, unlike the significant difference in PRV-1 expression observed on the mRNA level, the amount of PRV-1 protein on the cell surface is not consistently elevated in patients with polycythemia vera compared with healthy controls. Therefore, quantification of PRV-1 surface expression cannot be used for the diagnosis of polycythemia vera.
Klippel
Steffen
S
Department of Experimental Anaesthesiology, University Hospital Freiburg, Center for Clinical Research, and the Department of Hematology and Oncology, University Hospital Freiburg, Germany.
Strunck
Elisabeth
E
Busse
Christian E
CE
Behringer
Dirk
D
Pahl
Heike L
HL
D020413:3' Untranslated Regions / Q000235:genetics; D015139:Blotting, Southern; D002462:Cell Membrane / Q000502:physiology; D017931:DNA Primers; D058851:GPI-Linked Proteins; D006026:Glycoside Hydrolases; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000502:physiology*; D006801:Humans; D007519:Isoantigens; D008562:Membrane Glycoproteins; D011087:Polycythemia Vera / Q000097:blood* / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D015183:Restriction Mapping; D014158:Transcription, Genetic
D020413:3' Untranslated Regions; C407242:CD177 protein, human; D017931:DNA Primers; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols; D007519:Isoantigens; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D006026:Glycoside Hydrolases