The murine Cd48 gene: allelic polymorphism in the IgV‐like region
Authors: Cabrero J. G. , Freeman G. J. , Reiser H. 
Published: March 2003

The murine CD48 molecule is a member of the immunoglobulin superfamily which regulates the activation of T lymphocytes. Prior cloning experiments using mRNA from two different mouse strains had yielded discrepant sequences within the IgV‐like domain of murine CD48. To resolve this issue, we have directly sequenced genomic DNA of 10 laboratory strains and two inbred strains of wild origin. The results of our analysis reveal an allelic polymorphism within the IgV‐like domain of murine CD48.

The CD48 (Blast‐1, BCM1, sgp‐60) antigen is a cell surface molecule involved in the activation of murine T lymphocytes. Expression of CD48 is tightly controlled and restricted to lymphocytes, macrophages and dendritic cells; the molecule becomes upregulated upon cell activation. The counter‐receptor of CD48 is CD2 (T11) (Kato et al., 1992), the expression of which is similar to that of CD48 (Davis & van der Merwe, 1996). Antibodies to either CD2 or CD48 can block T‐cell activation in vitro (Arvieux et al., 1986; Reiser, 1990; Kato et al., 1992; Sen et al., 1992) and immune responses in vivo (Qin et al., 1994). Moreover, recent studies indicate that T cells from CD48‐deficient mice are defective in activation (our unpublished data).

CD48 is a member of the immunoglobulin (Ig) superfamily and, based on sequence homology and common chromosomal location, belongs to a subfamily of the Ig superfamily that – among other proteins – also includes its counter‐receptor CD2 (Davis & van der Merwe, 1996). The cDNA for murine CD48 has been isolated and is predicted to encode a protein composed of a signal peptide followed by an IgV‐like domain, an IgC‐like domain, and a membrane‐anchoring domain (Wong et al., 1990; Cabrero et al., 1993). Several findings have raised the possibility of an allelic polymorphism within the IgV‐like domain of murine CD48. Experiments in which CD48 was purified from a C57BL/6‐derived T‐cell line and subjected to microsequencing revealed the presence of an asparagine at residue 68 (Cabrero et al., 1993). The same amino acid was predicted from cDNA cloning experiments (codon AAT; nucleotides 318–320) in which lymphocyte mRNA from C57BL/6 mice was used as starting material for the construction of the library (Wong et al., 1990). In contrast, when CD48 was cloned from a BALB/c library the codon GAT was identified at the same position, predicting the presence of aspartic acid at residue 68 (Cabrero et al., 1993). Taken together these findings suggested the presence of an allelic polymorphism in the V‐like domain of murine CD48.

To further investigate this point we have directly sequenced genomic DNA from a total of 12 inbred strains. Of these strains, 10 were laboratory strains and two were inbred strains derived from wild origin. The results of the sequencing experiments are summarized in Fig. 1a. These data confirm that the Cd48 locus is characterized by an allelic polymorphism. Of the 10 laboratory strains examined, only C57BL/6J mice (accession number AF064620) carry codon AAT at position 318–320, whereas all other strains, including A/J (accession number AF064615), AKR/J (accession number AF064616), BALB/cJ (accession number AF064617), CBA/J (accession number AF064618), C3H/HeJ (accession number AF064619), DBA/1J (accession number AF064621), DBA/2J (accession number AF064622), SJL/J (accession number AF064623) and 129/SvJ (accession number AF064624) carry the codon GAT at this position. These data are consistent with prior cDNA and peptide sequence analysis and indicate that the CD48 protein derived from C57BL/6J mice carries an asparagine at amino acid residue 68 whereas the other laboratory strains carry aspartate at this position (Fig. 1b). Based on the results of molecular modelling studies by van der Merwe et al. (van der Merwe et al., 1995), this polymorphism is unlikely to affect binding of CD48 to its counter‐receptor CD2; whether it has other functional consequences remains to be determined.

Today’s classical laboratory strains represent a mosaic of various genetic components which are derived from the M. musculus complex. To further assess the polymorphism of the IgV‐like domain of Cd48 we included in our analysis genomic DNA from two inbred strains of wild origin, SKIVE/Ei and SPRET/Ei. These two strains are derived from Mus musculus and Mus spretus progenitors, respectively (Rex et al., 1987; Bonhomme & Guénet, 1996; Nakao et al., 1996). The latter subgenus is an evolutionarily distant relative of M. musculus and the standard laboratory strains (Bonhomme & Guénet, 1996). The Cd48 V‐like region of SKIVE/Ei was identical to that described above for BALB/c and other laboratory strains (Fig. 1a; accession number AF064625). In contrast, the V‐like region of Cd48 from SPRET/Ei mice differs from the laboratory strains at multiple positions, namely nucleotides 227, 248, 303, 317, 318, and 335 (Fig. 1a; accession number AF064626). Several of these substitutions are silent, but the substitutions of nucleotides 303, 317 and 318 lead to predicted amino acid changes at residues 63, 67 and 68 of the CD48 protein (Fig. 1b). These data are consistent with the evolutionary divergence between M. spretus on the one hand and M. musculus and the classical laboratory strains on the other.