ALTERATIONS IN REDOX BALANCE AND 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. Tucumán, Argentina.

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

We are interested in the physiological significance of the NADPH dependent P450 reductases expressed in T.cruzi. Increased resistance to classical trypanocidal compounds was previously demonstrated in TcCPR-B overexpressing T. cruziepimastigotes. Overexpression of TcCPR-A resulted to be lethal, showing abnormal cell shapes and altered DNA content. Taking together subcellular localization at ER and the biochemical behavior of recombinant TcCPR-C as Cyt P450 partner in reconstituted systems, we investigated its participation in lanosterol demethylation, a well described pathway catalized by CYP51, a cytochrome P450 family member in T. cruzi. Transgenic parasites as well as control epimastigotes were incubated with [3H]mevalonolactone, a precursor in 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 with or without induced oxidative stress by H2O2 treatment. Interestingly, TcCPR-C parasites present the most affected redox metabolism with a strong increase in NADP+/NADPH ratio, no longer modified by peroxides as it does occur in control parasites. NAD+/NADH ratio significatively decreased in transgenic lines compared to control.