Cathepsin D peptidase and a tyrosine phosphatase during follicular atresia in Dipetalogaster maxima (Hemiptera: Reduviidae): their involvement on early vitellin degradation

LEYRIA J; FRUTTERO LL; NAZAR M; CANAVOSO LE

Amsterdam

Congreso; The Seventh International Symposium on Molecular Insect Science; 2014

ELSEVIER

In insects, unfavorable nutritional conditions induce changes in the ovarian tissue which, in turn, promote follicular atresia. Throughout this process, early degradation of vitellin (Vt), the main yolk protein, and oocyte resorption take place. In the vectors of Chagas´disease, follicular atresia is poorly understood. In this context, the aim of this work was to analyze the biochemical and molecular events that take place during follicular atresia induced by nutritional deprivation in the hematophagous insect Dipetalogaster maxima, with special focus on cathepsin D (CD) and acid phosphatase (AP). For the study, fat bodies, ovaries and hemolymph were sampled at representative days of the reproductive cycle of the insect. The experimental approaches included enzymatic assays, immunofluorescence tests, western blot and RT-qPCR. In addition, in vitro assays of Vt proteolysis employing specific inhibitors were performed. The results showed that CD was expressed (mRNA and protein) at all reproductive stages in both, fat body and ovaries, being the expression levels of the active form of the enzyme significantly higher during atresia. In the hemolymph, only pro-cathepsin D was detected, being this pro-peptidase activated under optimal conditions. It was also shown that during the atretic stages, ovaries displayed the highest levels of AP and CD activities and that the main responsible for the AP activity was a tyrosine phosphatase (TyP). In vitro proteolysis assays demonstrated that AP and CD were necessary to prompt Vt degradation. By immunofluorescence we observed that CD and TyP were associated to yolk bodies of vitellogenic follicles. Altogether, our findings suggest that CD in the ovary has a dual origin (fat body and ovary) and that, together with a TyP, they are important in regulating early Vt proteolysis during follicular atresia either, to increase female lifespan or to sustain younger oocytes until improvement of nutritional conditions.