Reciprocal functions of the enzymes NBAD-synthase and NBAD-hydrolase in epidermal and neural tissue of flies

MARTÍN M. PÉREZ; ALEJANDRO RABOSSI; GERMAN SABIO; LUIS A QUESADA-ALLUÉ; ALEJANDRA BADARACCO

Buenos Aires

Congreso; Meeting of Teprhitid Workers of the Western Hemisphere; 2016

Tephritid Workers of the Western Hemisphere (TWWH)

The acylation of neurotransmitters is important for the homeostasis of neural system but also gives rise to important metabolites in insects. The best known are N-β-alanyldopamine (NBAD), the main sclerotization precursor of insect brown cuticles, and carcinine (N-β-alanylhistamine), an essential compound of the visual system. Key enzymes of this metabolism are NBAD-synthase (NBAD-S) and NBAD-hydrolase (NBAD-H) which synthesizes and hydrolyzes, respectively, β-alanyl derivatives of different neurotransmitters. These enzymes function together in a system regulating the levels of NBAD and dopamine during cuticle sclerotization, innate immune response and in nervous tissue. We studied the expression of NBAD-S and NBAD-H in different tissues during the life cycle of Ceratitis capitata and Drosophila melanogaster looking at protein expression visualized by western blots, immunohistochemistry and enzymatic activities. Synthesis and hydrolysis of beta-alanyl derivatives were determined using in vitro assays and analyzed by HPLC. Mutant flies were subjected to behavioral studies. We found that the two enzymes are expressed during the whole life cycle, from embryo to adult. NBAD-S is expressed periodically in epidermis induced by 20-0H-ecdysone only at the time of pupariation and of adult ecdysis, when the insect needs to sclerotize its cuticle. NBAD-H is active in a constitutive manner. Both enzymes are expressed constitutively in neural tissue. We characterized the melanic mutant niger of medfly Ceratitis capitata, lacking this enzymatic activity and demonstrated that this mutants show abnormal behavior and other deficiencies. We report the activity of these enzymes in different metabolic pathways and throughout the life cycle. Our results confirmed that NBAD-S and NBAD-H are essential during the development of insects and demonstrated that are necessary for the normal physiological balance of neural tissue.