Identification and expression analysis of a novel phytocystatin in developing and germinating seeds of triticale (×Triticosecale Wittm.)

In this paper the complete cDNA sequence of a newly identified triticale phytocystatin, TrcC-7, was analyzed. Because <em>TrcC-7</em> transcripts were present in seeds, we hypothesized that it may regulate storage protein accumulation and degradation. Therefore, changes in mRNA and protein levels during the entire period of seed development and germination were examined. Expression of <em>TrcC-7</em> increased during development and decreased at the end of maturation and subsequently increased during seed germination. Based on these results, TrcC-7 likely regulates cysteine proteinase activity during the accumulation and mobilization of storage proteins.

Cysteine proteinase activity is required for survival of the parasite in experimental acute amoebic liver abscesses in hamsters

Axenic trophozoites ofEntamoeba histolyticastrain HM1-IMSS grownin vitroin the presence of E-64, a potent irreversible inhibitor of cysteine proteinases, preserved their viability, their rate of replication, their resistance to complement, their haemolytic capacity and their ability to destroy target cells, despite complete inhibition of total cysteine proteinase activity. On the other hand, their erythrophagocytic capacity and their ability to decrease TER of MDCK cells was partially decreased. The same trophozoites injected into the portal vein of hamsters receiving a maintaining dose of E-64 failed to cause tissue damage and were rapidly eliminated. Our results suggest that amoebic cysteine proteinase activity is not required for amoebic functions inin vitroconditions, but that it becomes necessary for survival of trophozoites inin vivoconditions, whatever other role (if any) it may play in the parasite's virulence.

Toxopain-1 Is Critical for Infection in a Novel Chicken Embryo Model of Congenital Toxoplasmosis

ABSTRACT We tested the hypothesis that cathepsins and specifically toxopain-1, a cathepsin B, play a critical role in the pathogenesis of toxoplasmosis. We found that inhibiting the expression of toxopain-1-specific mRNA and protein by >60% significantly decreased the capacity of the parasites to multiply and invade in vitro. To relate these in vitro results to the role of toxopain-1 in pathogenesis in vivo, we developed a novel chicken embryo model of congenital toxoplasmosis. Inhibiting either toxopain-1 expression or specific cysteine proteinase activity significantly reduced congenital infection of chicken embryos, as determined by histopathology and by the number of parasites quantified by real-time PCR. Our new model provides key in vivo validation for the hypothesis that toxopain-1 is a potential drug target in Toxoplasma gondii and also provides a new animal model for rapid, inexpensive screening of antiparasitic compounds.