Experiment
Recombinant congenic strain AcB55 is resistant to malaria and is derived from parental strains A/J (susceptible) and C57BL/6J (resistant). An F2 cross between AcB55 and A/J was analyzed for linkage to malaria resistance. PklrChar4 on mouse Chromosome 3controls 30% of the phenotypic variance. A suggestive locus near D10Mit189 (7 cM) on mouse Chromosome 10 was further analyzed while controlling for PklrChar4. This resulted in significant linkage to malaria resistance at 4 cM near D10Mit167 (LOD=4.57).This locus controls 8% of the variance and is designated Char9 (P. chabaudi malaria resistance QTL 9). C57BL/6J-derived alleles at Char9 confer codominant malaria resistance.
The Char9 locus spans 14 Mb and contains 77 genes. RT-PCR analysis of A/J andAcB55 mRNA revealed highly significant differential expression of Vnn3, with AcB55 expressing 12-fold more Vnn3 mRNA compared to A/J (p<0.0001).
SNP analysis revealed a 4 Mb haplotype block conserved among 5 out of 6 known malaria-susceptible strains (SJL/J, C3H/HeJ, BALB/cJ, AKR/J, and A/J). Vnn3 maps to this haplotype block and displays 100% correlation between genotype and transcript expression level. The A/J-derived Vnn3 transcript carries a C1592T nucleotide substitution resulting in premature stop codon and 7 amino acid truncation.
In addition, Vnn3 and Vnn1, a similar gene 30 Kb downstream from Vnn3, are present in liver mRNA from C57BL/6 and AcB55 resistant strains but are absent in liver mRNA from susceptible A/J and AcB61 strains. Analysis of the Vnn3 putative proximal promoter region revealed the A/J sequence contains complex rearrangements that could possibly explain the lack of transcriptional activity in this strain.
Analysis Tools