Genetic and splice variations of Bos taurus CD46 shift cell permissivity to BVDV, the bovine pestivirus.

The pestivirus bovine viral diarrhea virus (BVDV) is known to bind to the CD46 molecule, which subsequently promotes entry of the virus. Mapping of the BVD-virion-binding site has shown that two peptides, 66EQIV69 and 82GQVLAL87, located on antiparallel beta sheets in the most distal complement control protein module (CCP1), provide the attachment platform. In the present study, we reveal the existence of ten distinct allelic versions of the CCP1 module, varying significantly in frequency among taurine and indicine races. A complex mRNA splicing pattern was also evidenced for bovine CD46, generating three different serine-threonine-proline segments and five different cytoplasmic domains. The four most frequent allelic variants and the six splice variants were then expressed in BVDV-nonpermissive porcine cells and the quantity of progeny virions generated by each cell preparation was measured 48 h post-infection. As expected, ectopic expression of the 10 bovine CD46 isoforms rendered the PK15 cells permissive to BVDV, as attested by the 100,000-fold greater recovery of virions from these cells than from non-transfected cells. This permissivity increase was significantly lower (-33%, P<0.001) when the canonical CCP1 was replaced with the variant most frequent in zebus, suggesting positive or negative selection of this allele in the latter and in the former, respectively. The predicted secondary structure of this variant suggests that the measured loss of function is due to the disappearance of one of the two beta sheets constituting the BVDV attachment platform. On the other hand we showed that for a given CCP1, the titer recovered at 48 hpi also depended on the nature of the CD46 cytoplasmic domain (P<0.001). This result implies that virus binding generates a cytoplasmic-tail-dependent outside-in signal that determines permissivity to BVDV.

Genomic structure, promoter analysis, and expression of the porcine (Sus scrofa) Mx1 gene.

Allelic polymorphisms at the mouse Mx1 locus affect the probability of survival after experimental influenzal disease, raising the possibility that marker-assisted selection using the homologous locus could improve the innate resistance of pigs to natural influenza infections. Several issues need to be resolved before efficient large scale screening of the allelic polymorphism at the porcine (Sus scrofa) Mx1 locus can be implemented. First, the Mx1 genomic structure has to be established and sufficient flanking intronic sequences have to be gathered to enable simple PCR amplification of the coding portions of the gene. Then, a basic knowledge of the promoter region needs to be obtained as an allelic variation there can significantly alter absolute levels and/or tissue-specificity of MX protein expression. The results gathered here show that the porcine Mx1 gene and promoter share the major structural and functional characteristics displayed by their homologs described in cattle, mouse, chicken, and man. The crucial function of the proximal interferon-sensitive response elements motif for gene expression is also demonstrated. The sequence data compiled here will allow an extensive analysis of the polymorphisms present among the widest spectrum possible of porcine breeds with the aim to identify an Mx1 allele providing antiviral resistance.