Identification And Function Exploration Of Ionotropic Receptors And Pheromone Binding Protein In Locust

Two novel ionotropic receptors, IR8a and IR25a were firstly identified in grasshopper, Schistocerca gregaria. Further research was conducted in spatiotemporal expression pattern, cell localization and so on. A putative courtship related pheromone was defined and a possible pheromone binding protein-OBP4may be involved in the process of sensing this pheromone, details are as follows,I. Identification and Characterization of IR8a and IR25a in locust, Schistocerca gregariaIn this study, we have identified the genes of S. gregaria which encode homologues of co-receptors for the variant ionotropic receptors, the subtypes IR8a and IR25a. It was found that both subtypes, SgreIR8a and SgreIR25a, was expressed in the antennae of all five nymphal stages and adults. Attempts to assign the relevant cell types by means of in situ hybridization revealed that SgreIR8a and SgreIR25a are expressed in cells of sensilla coeloconica. Double fluorescence in situ hybridization experiments disclosed that the two IR-subtypes are co-expressed in some cells of the coeloconic sensilla. Expression of SgreIR25a was also found in some of the sensilla chaetica, however, neither SgreIR25a nor SgreIRSa was found to be expressed in basiconic or trichoid sensilla. This observation was substantiated by the results of double FISH experiments demonstrating that cells expressing ISgreIR8a or SgreIB25a do not express SgreORco. These results support the notion that the antenna of the desert locust employs two different populations of OSNs to sense odors:cells which express IRs in sensilla coeloconica and cells which express ORs in sensilla basiconica and sensilla trichodea.II. A dual-role male pheromone sensed by odorant-binding protein in locustHere for the first time, we identified KT as one courtship related pheromoene, a component of male adult body volatiles of locust, Locusta migratoria. KT has been reported by Cui et al., that can excite or inhibit neurons in a portion of sensilla trichodea. This compound elicited high electrophysiological responses of male and female antenna. Behavioural assays showed that KT evoked sexually dimorphic behaviours at certain dosage, attracting females and repelling males but5instar nymphs did not show any response to KT. Together these results suggested that this compound is likely to be one courtship related pheromone of L. migratoria. Consequently, this pheromone was demonstrated to have binding affinity to a novel odorant binding protein which was identified from locust, named as LmigOBP4, highly expressed in olfactory sensilla observed in in situ hybridyzation and immunocytochemistry localization experiments. Furthermore, the defect of LmigOBP4by RNAi technique diminished significantly electroantennogram responses of both male and female locusts to KT. Consistently, behavioural tendencies in the selection of KT at certain doses by both sexes were reversed with that of the comparison of wild types. Additionally, behavioural impairment of KT by RNA interference of Lmigobp4could not be restored and compensated by increasing doses. Therefore, we have characterized a dual-role locust male-specific pheromone, KT which can elicit courtship behaviour, attracting females and repelling males, and the pheromone is sensed by a novel odorant binding protein, LmigOBP4. Our findings support that odorant binding protein plays key roles in the process of pheromone detection, and locust has highly efficient courtship communication system which is more similar to fly, other than moth. This provides the evidence to understand the molecular evolution of animal courtship, as well as to develop new methods for controling locust plagues based on functions of molecules in locust courtship behavior.