Fig. 6.
Chaperone activity of mTXNPxs is critical for Leishmania virulence. (A) Influence of a 20-fold molar excess of His.THR-mTXNPxred on the in vitro thermal aggregation of luciferase. See the legend of Fig. 1 for details. (B) Analysis of mTXNPx expression in mtxnpx− (lane 1), mtxnpx−/+mTXNPx (lane 2), and two mtxnpx−/+MTS.His.THR-mTXNPx (lanes 3 and 4, respectively) promastigote clones using a polyclonal antibody raised against the recombinant His.THR-MTS.mTXNPx. Purified mTXNPx and His.THR-mTXNPx were used as controls for the expected size of the proteins, and their position on the blot is indicated. Ponceau S staining of the blot is shown as loading control. (C) Localization of the His.THR-mTXNPx chimera in mtxnpx− promastigotes using indirect immunofluorescence with the anti-mTXNPx antibody (green) merged with DAPI (blue). Controls as described above are included. (D) Growth rate of mtxnpx− (black circles), mtxnpx−/+mTXNPx (black squares), and two mtxnpx−/+MTS.His.THR-mTXNPx clones (dark gray triangles). (E) Parasite burden of BALB/c mice after 2 and 8 wk of infection. BALB/c mice were inoculated i.p. with stationary-phase promastigotes of the mtxnpx−/+mTXNPx clone (squares) or two mtxnpx−/+MTS.His.THR-mTXNPx clones (triangles). After 2 and 8 wk of infection, the number of parasites per gram of spleen (parasite burden, PB) was determined. Each point represents one animal. The red line indicates the detection limit of the technique (log10 = 2.7). Animals with parasite burdens below this limit are “not detected” (ND) and are represented by red symbols.