Biochemical and Biophysical Research Communications
The murine pan T cell marker CD96 is an adhesion receptor for CD155 and nectin-1
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Cloning of mCD96; cloning, expression, and purification of recombinant mCD96-hIgG1, mNectin-1/2/3-hIgG1, and mCD226-hIgG1
mCD96 was cloned using primers MUCD96.BAM and mucd96_full in PCR with cDNA from BALB/c spleen (MTC panel, CLONTECH): 2 min 94 °C, 25 cycles with 15 s 94 °C, 30 s 55 °C, 1 min 72 °C. The PCR fragment, cloned into pcDNA3, served as template in a subsequent PCR applying primers MUC96.BAM and mucd96_fus_bam to generate an ectodomain fragment for in frame cloning into vector phIgG1 (containing the cDNA for domains 3 + 4 of hIgG1).
PCR products representing the ectodomains of mNectin-1, mNectin-2, mNectin-3,
Cloning of mCD96 and binding to mCD155
Using splenic cDNA from BALB/c mice, a full-length clone spanning the putative mCD96 open-reading frame was generated by PCR. The obtained product was sequenced and found to be identical to the published sequence (Accession No. Y029156). An alignment of the human and murine protein sequence reveals 56% conservation at the amino acid level (Supplementary Fig. S1).
Apart from CD226, CD155 binds to CD96 expressed on human NK cells [4]. To investigate whether the CD155/CD96 interaction is conserved
Acknowledgments
This work was funded by DFG-Grant BE1886/2-1 from Deutsche Forschungsgemeinschaft to G. Bernhardt. S. Seth is a recipient of a Georg Christoph Lichtenberg Stipend provided by the Ph.D. program Infection Biology sponsored by the Ministry of Science and Culture of Lower Saxony.
We thank O. Pabst for critically reading the manuscript.
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These authors contributed equally to this work.