Alterations in redox balance ande ergosterol synthesis in TcCPR overexpressing T. cruzi epimastigotes

PORTAL P; DE VAS MG; FLAWIA MM; TORRES HN; ALONSO GD; PAVETO C

San Miguel de Tucumán -Argentina

Congreso; XLV Reunión anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB),; 2009

SAIB

Increased resistance to classical trypanocidal compounds was previously demonstrated in the NADPH dependent reductase TcCPR-B overexpressing T.cruzi epimastigote cells. Overexpression of TcCPR-A produced abnormal cell shapes and cell division with cultured death upon selection. Taking together subcellular localization at ER and the biochemical behavior of purified recombinant TcCPR-C as cytochrome P450 partner in reconstituted systems, we decided to investigate its participation in lanosterol demethylation, a well described pathway catalized by CYP51, a cytochrome P450 family member in T.cruzi. Transgenics parasites as well as control epimastigotes were incubated with [3H]mevalonolactone, a precursor of ergosterol, the characteristic sterol in trypanosomatids and yeast cells. An increase in ergosterol content could be observed in overexpressing TcCPR-C epimastigotes. Redox balance (NADP+/NADPH and NAD+/NADH ratios) was also investigated in the transgenic lines with or without induced oxidative stress by H2O2 treatment. Interestingly, TcCPR-C parasites present the most affected redox metabolism showing a strong increase in NADP+/NADPH ratio, which is no longer modified by peroxides as it does occur in control parasites. NAD+/NADH ratio significatively decreased in transgenic line compared to control while total intracellular concentration of pyridine nucleotides remained unchanged.