DmCatD, a cathepsin D-like peptidase of the hematophagous insect vector Dipetalogaster maxima (Hemiptera: Reduviidae): bioinformatic analysis and internalization pathway in developing oocytes

SETTEMBRINI BEATRIZ P; FRUTTERO LL; LEYRIA J; CANAVOSO LE

Orlando

Congreso; XXIV International Congress of Entomology; 2016

Entomological Society of America

In this work, we have investigated the involvement of cathepsin D and acid phosphatase in the process offollicular atresia of Dipetalogastermaxima, a hematophagous insect vector of Chagas? disease. For the studies, fat bodies, ovaries and hemolymph were sampled fromanautogenous females at representative days of the reproductive cycle:pre-vitellogenesis, vitellogenesis as well as early and late atresia. Real time PCR (qPCR) and western blot assays showed that cathepsin D wasexpressed in fat bodies and ovaries in all reproductive stages, being theexpression of its active form significantly higher at the atretic stages. In thehemolymph, only the immunoreactive band compatible with pro-cathepsin D wasobserved by western blot. On the other hand, acid phosphatase activity inovarian tissues significantly increased during follicular atresia in comparisonto pre-vitellogenesis and vitellogenesis. A further enzyme characterization with inhibitors showedthat the high levels of acid phosphatase activity in atretic ovaries corresponded mainly to a tyrosinephosphatase. In vitellogenic ovarian follicles, cathepsin D and tyrosine phosphatase were localized interminal oocytes, associated with yolk granules. It was also shown that bothenzymes partially co-localized with vitellin. In early atresia however, thedistribution pattern of cathepsin D and tyrosine phosphatase changed noticeably.In vitro assays using homogenates of atreticovaries as the enzyme source and enzyme inhibitors demonstrated that cathepsin D, together with a tyrosinephosphatase, were necessary to promote thedegradation of vitellin. Taken together, the results strongly suggested thatboth acid hydrolases play a central role in early proteolysis of vitellinduring the process of follicular atresia.