Biological Analysis Of Ribosomal Protein Family And MAPK Singaling Pathway Associated Genes In Apis Cerana Cerana

Insects undergo multiple morphological changes, including organ formation, tissue mature,throughout the process of development, and these organism changes are regulated by manygenes expression. Extensive studies have elucidated some of the genes, such as the ribosomalprotein (RP) family genes and those in the mitogen activated protein kinase (MAPK) pathway,that are required for development. So far, researches about ribosomal protein and MAPKsingaling pathway associated genes are mainly focus on the model speices, such asArabidopsis thaliana, Homo sapiens and Mus musculus, and information in honeybee isrelatively limited. The Asian honeybee (Apis cerana cerana) is an important indigenousspecies and has important roles in the pollination of flowing crops. In this study, we selectedthe economically important species A. cerana cerana as the experiment material, and threegenes, AccRPL17, AccRPL11, AccTAK1, have been cloned by RT-PCR and RACE-PCR.Then, a series of studies have been conducted on the sequence and expression analysis,function prediction of three genes. The main results are as follows:(1) Alignment analysis revealed that AccRPL17, AccRPL11and AccTAK1shared82.07%,83.46%and63.27%similarities with other known species. Conserved characteristicregions were identified in their amino acid sequences, respectively.(2) Through the TFSEARCH database, several transcription factor binding sites relatedto early development were identified in the5’-flanking region of AccRPL17and AccTAK1,such as DRI, CRE, BR-C. A binding motif for Croc, a brain development-relatedtranscriptional activator, was also found in AccRPL11promoter region. These results suggestthat AccRPL17and AccTAK1maybe related to early development, and AccRPL11is likely tofulfill a function in brain maturation during adult phase.(3) qRT-PCR and Northern blot were performed to examine the developmentalregulation and tissue distribution of AccRPL17, AccRPL11and AccTAK1. qRT-PCR showed that the highest mRNA level of AccRPL17was detected in larvae on the fifth day. AccTAK1was expressed at high levels in fourth instar larvae, primarily in the midgut and salivaryglands. These results suggest that AccRPL17might play an important role in insectdevelopment, and AccTAK1may be involved in the regulation of early development in thelarval salivary gland and midgut. Northern blot revealed that AccRPL11expression washighly concentrated in the adult brain, and no significant difference in AccRPL11expressionbetween males and females. The results suggest that AccRPL11may be function in brainmaturation during honeybee adults.(4) qRT-PCR showed that the transcript levels of AccRPL17, AccRPL11and AccTAK1can be markedly accumulated by treatment with various abiotic stimulis such as pesticides,heavy metals and organic solvents. Those results indicate that AccRPL17, AccRPL11andAccTAK1may be involved in responses to abiotic stresses, and play an important role intissues development.(5) One-day post-emergence adults were injected with dsAccRPL11. Northern blot andELISA displayed that lower transcript levels were observed in the dsAccRPL11-injected adultsat5and9days post-injection. Therefore, we selected adults at7days post-injection to furtheranalyze AccRPL11expression in brain. Q-PCR experiments showed that the expression levelof AccRPL11in7-day post-injection adults was more severely decreased in the brain than inother tissues. RT-PCR showed that the expression levels of four other braindevelopment-related genes, p38, ERK2, CacyBP and CREB, were also reduced indsAccRPL11-injected adults. These results suggest that the inhibition mediated bydsAccRPL11is effective in adult brain, and AccRPL11may be function in brain maturationduring adult phase.(6) Immunohistochemical localization showed that AccRPL17is primarily concentratedin muscular tissues, stigma, body wall and the surrounding of the eye in the fifth instar larvae.These results suggest that AccRPL17may play an important role in early development.AccTAK1was expressed primarily in the intestinal wall cells of the midgut and the secretorycells of the salivary glands in fourth instar larvae. These results indicate that AccTAK1maybe involved in the regulation of early development in the larval salivary gland and midgut.AccRPL11was localized to the medulla, lobula and surrounding tissues of esophagus in the brain. However, immunofluorescence signal attenuation was observed in AccRPL11-richregions of the brain in dsAccRPL11-injected honeybees. These results suggest that AccRPL11may be function in brain maturation during honeybee adults.