A study of the coregulation and tissue specificity of XG and MIC2 gene expression in eukaryotic cells.
CD99, the product of the MIC2 gene, exhibits an erythroid-specific quantitative polymorphism coregulated with the polymorphism of the XG blood group gene. As a preliminary study of this phenomenon, human XG and CD99 recombinant proteins were expressed in murine RAG cells and analyzed by flow cytometry. Both proteins were expressed independently and at a similar level in single and double transfectants. Immunoprecipitation and Western blot analysis, using the murine monoclonal antibodies NBL-1 and 12E7, revealed species of 26 kd (XG) and 32 kd (CD99), respectively. A putative 28-kd intracellular precursor of CD99 was also detected, as was a 26-kd species after neuraminidase treatment of CD99-expressing cells. No evidence of association or complex formation between XG and CD99 proteins could be proven, either on transfected RAG cells or on human erythrocytes. These results were confirmed using somatic hybrids between single transfectants. These findings suggest that the phenotypic relationship between XG and CD99 is mostly regulated at the transcriptional level, but they do not formally exclude some posttranscriptional effect. Studies on the tissue specificity of XG expression showed that surface expression of the XG protein could not be restored in somatic hybrids between B-lymphoblastoid cell lines from Xg(a+) persons and fibroblasts (RAG) or erythroid (MEL) cells. RT-PCR analysis of the transcripts revealed the existence of an XG mRNA in each cell line, suggesting that the tissue-specific regulation of cell surface XG expression occurs either at a quantitative transcriptional level or is a posttranscriptional event. By Northern blot analysis, XG transcripts were detected in erythroid tissues and several nonerythroid tissues. (Blood. 2000;95:1819-1826)
Fouchet
C
C
Inserm U76, Institut National de la Transfusion Sanguine, Paris, France.
Gane
P
P
Huet
M
M
Fellous
M
M
Rouger
P
P
Banting
G
G
Cartron
J P
JP
Lopez
C
C
D000071443:12E7 Antigen; D000230:Adenocarcinoma / Q000473:pathology; D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis* / Q000235:genetics; D001789:Blood Group Antigens; D015815:Cell Adhesion Molecules / Q000096:biosynthesis* / Q000235:genetics; D004912:Erythrocytes / Q000378:metabolism; D005057:Eukaryotic Cells / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D006822:Hybrid Cells / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D009928:Organ Specificity; D011498:Protein Precursors / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D000071443:12E7 Antigen; D015703:Antigens, CD; D001789:Blood Group Antigens; C061828:CD99 protein, human; D015815:Cell Adhesion Molecules; D009363:Neoplasm Proteins; D011498:Protein Precursors; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C088785:XG protein, human