Abstract
Calmodulin–ubiquitin ( cub) is a single-copy gene of Trypanosoma cruzi, which encodes a 208 aminoacid polypeptide of unknown function, containing putative calcium-binding domains. After targeted deletion, a clone (TulCub8) was derived where one of the two alleles was disrupted. This clone displayed a sharp and stable loss of virulence for mice. Parasitemias after inoculation of 10 6 trypomastigotes of the mutant, as compared to wild-type parasites were 68-fold lower ( p = 0.018) in adult Swiss mice and 27-fold lower ( p = 0.002) in newborn Balb/c mice. Epimastigote inocula of the mutant were strongly protective against infection by wild-type parasites. Virulence was not restored by serial passage in mice, showing that the attenuated phenotype is stable and gene-conversion from the intact cub allele does not occur at an appreciable rate. Retransfection of the missing cub allele restored virulence. Complementation experiments showed that the intact cub gene is necessary for full expression of virulence.
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