A polycistronic retrovirus vector for expression of swine MHC class II DRα/β heterodimers


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Abstract

A recombinant polycistronic retroviral vector was generated to express multimeric proteins from a single transcript by using internal ribosomal entry sites which allow independent initiation of translation from internal cistrons. In addition, an enhancerless SV40 origin of replication was incorporated into the recombinant plasmids between the retroviral long terminal repeat sequences to facilitate the testing of expression of multiple proteins in a transient COS cell transfection assay.With the ultimate goal of utilizing these vector types for the induction of transplantation tolerance via retrovirus-mediated gene transfer (LeGuern et al. J Mol Med 1995:73:269[1]; Sachs Transplant Sci 1993:3:59[2] ) we have incorporated miniature swine MHC class II DRA and DRB cDNAs which encode for the SLA-DRα and SLA-DRβ polypeptide chains, respectively, into the retroviral construct. The proviral DNA integrated within the producer clone genome, was found to be stable over a 3-month period and no loss of expression of DR heterodimers was observed on the cell surface of either the retrovirus producer cells or the retrovirally transduced fibroblast cell line. High titers (>1 x 106 CFU/ml) of recombinant particles, devoid of replication competent helper viruses, were obtained. Such vectors represent the first successful attempt to express multimeric proteins at the cell surface of transduced cells.

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