The APO-1/Fas (CD95) receptor is expressed in homozygous MRL/lpr mice.
APO-1/Fas (CD95) is a transmembrane receptor that transduces apoptotic signals within various cells including T and B cells. The APO-1 gene was found to be defective in lpr mice. In these mice insertion of a retrotransposon gives rise to transcription of an abnormal mRNA and only a fraction of wild-type APO-1 mRNA. It is not clear if lpr mice still express wild-type APO-1 protein. To address this question, we prepared rabbit anti-APO-1 antibodies (Ab) with a peptide representing the extracellular sequence corresponding to residues 5-23 of APO-1. The rabbit Ab reacted with thymocytes from different mouse strains and the extent of binding was correlated with the two known APO-1 alleles. In addition, the Ab reacted with mouse cell lines expressing mouse APO-1 mRNA but not with human cell lines. Binding of the Ab to MRL and BALB/c thymocytes was completely blocked by the immunizing peptide. Immunofluorescence analysis of MRL/lpr thymocytes showed that they still express APO-1 protein at approximately one tenth of the wild-type expression level on their surface. In addition, in lpr as in wild-type mice we found a decrease of APO-1 expression in the more mature thymic compartment. Western blot analysis of whole cell lysates from lpr and wild-type thymocytes showed that the Ab recognized APO-1 in both cell types. Approximately 50% of CD3+ splenocytes and 80% of in vitro activated CD3+ cells from wild-type mice reacted with the Ab, but to a lower extent than thymocytes. The same differential reactivity was found in lpr CD3+ splenocytes. lpr T cells, however, showed a substantially lower level of APO-1. Thus, the differential expression of APO-1 on thymic versus peripheral lpr T cells might influence their sensitivity towards APO-1-mediated apoptosis.
Mariani
S M
SM
Division of Immunogenetics, German Cancer Research Center, Heidelberg.
Matiba
B
B
Armandola
E A
EA
Krammer
P H
PH
D000483:Alleles; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D000954:Antigens, Surface / Q000032:analysis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D017209:Apoptosis; D001483:Base Sequence; D017252:CD3 Complex / Q000032:analysis; D017931:DNA Primers / Q000737:chemistry; D005434:Flow Cytometry; D015870:Gene Expression; D017493:Leukocyte Common Antigens; D051379:Mice; D008815:Mice, Inbred Strains; D008817:Mice, Mutant Strains; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D013601:T-Lymphocytes / Q000378:metabolism; D013950:Thymus Gland / Q000166:cytology / Q000378:metabolism; D019014:fas Receptor
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D017252:CD3 Complex; D017931:DNA Primers; D012333:RNA, Messenger; D019014:fas Receptor; D017493:Leukocyte Common Antigens