| In recent years,the shellfish aquaculture industry in China has been plagued by frequent outbreaks of diseases,posing a serious threat to its sustainable development.Bacterial and viral infections are among the main causes of mass mortality in shellfish.The scallop species Zhikong scallop(Chlamys farreri)is one of the important cultured varieties,which lacks specific immunity and relies only on innate immunity to combat pathogen infections.Therefore,elucidating the immune mechanisms underlying the disease resistance in Chlamys farreri is of great significance for the development of disease prevention and control strategies.This study focuses on a key factor in the immune signaling pathway of Chlamys farreri,the inhibitor of nuclear factor kappa-B kinase(IKK),and investigates the specific role of the IKK gene in the natural immune processes.The main research objectives are as follows:1.In this study,we cloned and identified the classical IκB kinase family genes,CfIKK1 and CfNEMO,in the scallop species Chlamys farreri.We analyzed their protein domain composition and determined their specific classification status.At the transcriptional level,qRT-PCR results revealed that CfIKK1 and CfNEMO were widely expressed in all examined tissues.CfIKK1 showed the highest expression in the hepatopancreas,while CfNEMO exhibited high expression in the gill tissue.Furthermore,both CfIKK1 and CfNEMO genes were responsive to different pathogen-associated molecular patterns(PAMPs)stimuli.Furthermore,the results of the Co-Immunoprecipitation(Co-IP)experiment indicate that the CfIKK1 protein interacts specifically with Myeloid differentiation factor 88,(MyD88)and participates in Toll-like receptor(TLR)signaling transduction pathways.The CfIKK1 protein forms homodimers through self-binding and may further associate with CfNEMO to form the IKK complex.RNA interferenceexperiments demonstrate that inhibiting the expression of the CfIKK1 gene leads to reduced expression of CfIRFs genes.These experimental findings suggest that CfIKK1 and CfNEMO play crucial roles as signaling molecules and immune regulatory factors in the innate immune system of scallops.2.The CfIKK2 gene,a member of the non-classical IκB kinase family,has been cloned and identified in the scallop species known as Chlamys farreri.The CfIKK2 gene is widely expressed in various tissues,with higher expression levels observed in the liver and pancreas.Its expression is significantly influenced by Poly(I:C)and acute virus necrobiotic virus(AVNV)stressors.The CfIKK2 protein interacts with the MyD88 protein through its CCD1 domain,participating in the signal transduction of TLR pathways.The kinase domain(KD)and CCD1 domain of CfIKK2 protein facilitate the formation of homodimers.Overexpression of CfIKK2 protein can significantly activate interferons(IFNs),suggesting that the CfIKK2 gene may exert immune effects through IFNs activation.In conclusion,this study has provided a preliminary analysis of the role of the IKK gene in the natural immune defense process of the scallop Chlamys farreri,elucidating the immune pathway signaling mechanism mediated by the IκB kinase in scallops.The research findings have enriched the theoretical understanding of the natural immune system in scallops and have laid a solid theoretical foundation for disease prevention and control as well as the selection and breeding of disease-resistant strains in the aquaculture industry. |
