Submitted to: Insect Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/25/2013
Publication Date: 3/28/2013
Citation: Nusawardani, T., Kroemer, J.A., Choi, M.Y., Jurenka, R.A. 2013. Identification and characterization of the pyrokinin/PBAN family of GPCRs from Ostrinia nubilalis. Insect Molecular Biology. 22(3):331-340. Interpretive Summary: Lepidopteran insects are major pests of agriculture throughout the world, and their control is responsible for the largest use of chemical pesticides. Novel biologically-based or environment friendly methods of controlling these pests are needed to reduce our dependence on pesticides. Insect neuropeptides play many critical roles in insect development and metamorphosis. Identifying and understanding the mode of action for these neuropeptides and their receptors could lead to new non-insecticidal control methods. Scientists at the Center for Medical, Agricultural and Veterinary Entomology, USDA, ARS, Gainesville, FL, and the Department of Entomology, Iowa State University, Ames, Iowa, have characterized three membrane-bound receptors, including PBAN (pheromone biosynthesis activating neuropeptide)-receptor; Diapause hormone (DH) receptor; and a Periviscerokinin (PVK) receptor from the European corn borer. The PBAN-R, DH-R and PVK-R are G-Protein coupled receptors (GPCR) consisting of 7-transmembrane domains that form three (3) extracellular loops on the cellular surface. In the present study, we identified and characterized three receptors and functionally expressed them in an insect cell line for binding studies with peptide ligands. The PBAN-R gene sequence has been identified in about a dozen lepidopteran species. In contrast the DH-R has only been characterized in the silkworm . Thus, this report describes the characterization of all three closely related receptors in a single lepidopteran species for the first time.
Technical Abstract: Neuropeptides regulate a variety of physiological functions in most animals. A major groups of insect neuropeptide hormones is the pyrokinin (PK) /PBAN (Pheromone Biosynthesis Activating Neuropeptide) family. Insects have at least two closely related G-protein coupled receptors (GPCR) for the family, one binding PBAN and another binding diapause hormone (DH). A third related receptor binds a product of the capa gene, periviscerokinins. Here we identify these three receptors from the European corn borer, Ostrinia nubilalis. Quantitative PCR of the receptor mRNA indicates differential expression in various life stages and tissues, which will be compared with findings in other insects. All three splice variants of the PBAN-receptor were identified with all variants found in pheromone gland tissue. An immunofluorescence expression assay indicates that all three variants can be expressed to the cell surface of Sf9 cells. The A- and B-variants were not active in our functional assay, which confirms studies in other moths. Functional expression of the PBAN-rC variant indicates that it is the most active with a 44 nM half effective concentration (EC50) for activation by PBAN. The DH-receptor was activated by DH with an EC50 value of 150 nM.