The role of LuloPAT amino acid/proton symporters in midgut alkalinization in the sandfly Lutzomyia longipalpis (Diptera - Psychodidae).

ABSTRACT

In Lutzomyia longipalpis females, which are the main vectors of Leishmania infantum in the Americas, hematophagy is crucial for ovary development. The control of pH in the midgut during blood digestion is important to the functioning of the digestive enzymes, which release amino acids in the luminal compartment that are then transported through the enterocytes to the hemolymph for delivery to the ovary and other organs. In the present work, we investigated transport systems known as LuloPATs that are present in the midgut of L. longipalpis but not in other organs. These transporters achieve symport of amino acids with H+ ions, and one of them (LuloPAT1) is orthologous to a transporter described in Aedes aegypti. According to our results, the transcription levels of LuloPAT1 increased significantly immediately after a blood meal. Based on the variation of the fluorescence of fluorescein with the pH of the medium, we developed a technique that shows the acidification of the cytoplasm of gut cells when amino acids are cotransported with H+ from the lumen into the enterocytes. In our experiments, the midguts of the sandflies were dissected and opened longitudinally so that added amino acids could enter the enterocytes via the lumen (PAT carriers are apical). LuloPAT1 transporters are part of a complex of mechanisms that act synergistically to promote gut alkalinization as soon as blood intake by the vector occurs. In dissected but not longitudinally opened midguts, added amino acids could only enter through the basolateral region of enterocytes. However, alkalinization of the lumen was observed because the entry of some amino acids into the cytoplasm of enterocytes triggers a luminal alkalinization mechanism independent of LuloPATs. These findings provide new perspectives that will enable the characterization of the set of signaling pathways involved in pH regulation within the L. longipalpis midgut.