Function Characteristics Of Three Neuropeptide Genes Nl18169, Nl35689 And Nl35441 In Nilaparvata Lugens

Insect neuropeptides play key roles in regulation of metabolism, development, reproduction, rhythm, diapause, behavior and learning. Deciphering the molecular basis of their regulation will provide new insights into the mechanisms of insect growth, development and behavior as well as provide knowledge for exploiting new control methods of insect pests. Brown planthopper (BPH), Nilaparvata lugens (Homoptera:Delphacidae), is one of the most important rice insect pests. However, the functions of neuropeptides in BPH remain largely unknown. In this study, we cloned three neuropeptide genes, one crustacean cardioactive peptide (CCAP) gene Nl18169 and two bursicon genes N135689 and N135441, in BPH and investigated their functions in growth, development, survival, feeding and virulence change in BPH.CCAPs are important neuropeptides in insects and they have multiple physiological roles, such as controlling heart rate and modulating ecdysis behaviour. Here we cloned a BPH CCAP gene Nl18169 with an open reading frame (ORF) of 408bp. The amino acid sequence deduced from the ORF revealed that Nl18169 encodes a protein of 135 amino acids (Fig.1) with a calculated Mr of 15.2kDa and a theoretical pI of 8.36. The protein N118169 possesses a singal peptide with 24 amino acids but there was no transmembrane domain. RT-PCR analysis showed that Nl18169 was highly expressed in the head and salivary-gland and more highly expressed in Mudgo population than in TN1 population. Knockdown of N118169 decreased the survival rate and influenced molting and eclosion of BPHs but did not cause a difference in survival rate and feeding when BPHs were placed on rice varieties with different levels of resistance (Mudgo and TN1), indicating that N118169 is crucial in development and survival of BPH but not important in virulence change.Bursicons also belong to neuropeptides in insects, and are involved in the process of insect eclosion. We cloned two BPH bursicon genes, called N135689 and N135441. The cDNA of N135689 is 566bp with an ORF of 462bp. The amino acid sequence deduced from the ORF revealed that N135689 encodes a protein of 154 amino acids with a calculated Mr of 17.2kDa and a theoretical pI of 6.87. N135441,540bp in full-length with an ORF of 417bp, encodes a protein of 139 amino acids with a calculated Mr of 15.7kDa and a theoretical pI of 5.07. Both N135689 and N135441 have no transmembrane domain but each possesses a signal peptide, with 21 and 24 amino acids, respectively. RT-PCR analysis revealed that N135689 and N135441 were highly expressed in the head and there was no difference in mRNA levels of both genes between two BPH populations, Mudgo population and TN1 population. Knockdown of Nl35689 or Nl35441 also decreased the survival rate of BPH and lead to abnormality in molting and eclosion. The results indicate that Nl35689 and Nl35441 are important in the development and survival of BPH.