RNA-Seq Analysis Revealed Differential Gene Expression and Exon-Specific Effects Associated with the Mating Process in Honey Bee Queens

Document Type : Original Research

Authors
S. Alipour
A. A. Masoudi
Department of Animal Science, Faculty of Agriculture, Tarbiat Modares University, Tehran, Islamic Republic of Iran.
10.22034/jast.25.4.7
Abstract
Mating in honeybees causes dramatic changes not only in its behavior and physiology but in genes’ transcriptional level. To determine the molecular mechanisms regulating post-mating behavioral changes, we examined gene expression modification and exon-specific expression of the virgin queen versus the queen injected with semen in hemocoel. The DESeq2 package of R was used to identify the Differentially Expressed Genes (DEGs). The DEGs were selected for functional enrichment analysis and Protein-Protein Interaction (PPI) network construction. We also performed differential exon usage analysis using the DEXseq R package. Results identified a significant expression (FDR< 0.05) of a total of 971 genes between two groups of insects. The mating process produced significant changes in the expression of cell surface receptor signaling pathway, innate immune response, extracellular region, proteinaceous extracellular matrix, nucleous, G-protein coupled receptor activity, heme binding, and transmembrane transporter activity genes. Protein-Protein Interaction (PPI) network shows that LOC552504 (titin-like) could be considered as a super-hub gene in the mating process of queens. In addition, we identified exons that were differentially expressed in two groups of honeybee queens. At 10% FDR, we found significant differential exon usage in 79 genes. Among them, GB55396 gene had the most differences in exon usage and could be the best candidate gene for mating and reproductive activation in queens.

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Albert, R., Jeong, H. and Barabasi, A.L. 2000. Error and attack tolerance of complex networks. Nature, 406: 378–82.
Anders, S., Reyes, A. and Huber, W. 2012. Detecting differential usage of exons from RNA-seq data. Genome Research, 22(10): 2008–17.
Andrews S. 2010. FastQC: a quality control tool for high throughput sequence data. http://www.bioinformatics.babraham.ac.uk/projects/fastqc/
Baumann, H. 1974. Biological efects of paragonial substances PS-1 and PS-2, in females of Drosophila funebris. Journal of Insect Physiology, 20: 2347–2362.
Berger, B., Abdalla, F.C. and Cruz-Landim, C. 2005. Effect of narcosis with CO2 on the ovarian development in queens of Apis mellifera (Hymenoptera, Apini). Sociobiology, 261–270.
Black, D.L. 2003. Mechanisms of alternative pre-messenger RNA splicing. Annual review of biochemistry, 72: 291–336.
Dennis, G., Sherman, B.T., Hosack, D.A., Yang, J., Gao, W., Lane, H.C. and Lempicki, R.A. 2003. DAVID: database for annotation, visualization, and integrated discovery. Genome Biology, 4: R60.
Ebadi, R., Gary, N.E. and Lorenzen, K. 1980. Effects of carbon dioxide and low temperature narcosis on honey bees, Apis mellifera. Environmental Entomology, 9(1): 144–8.
Gary, N. 1992. Graham JM, ed. The hive and the honey bee. Hamilton: Dadant & Sons, Inc, 271–307.
Harris, J.W., Woodring, J. and Harbo, J.R. 1996. Effects of carbon dioxide on levels of biogenic amines in the brains of queenless worker and virgin queen honey bees (Apis mellifera). Journal of Apicultural Research, 35(2): 69–78.
Harris, J.W., Woodring, J. and Harbo, J.R. 1996. Effects of carbon dioxide on levels of biogenic amines in the brains of queenless worker and virgin queen honey bees (Apis mellifera). Journal of Apicultural Research, 35(2): 69–78.
Harris, R.M. and Hofmann, H.A. 2014. Neurogenomics of Behavioral Plasticity. In: Ecological Genomics. Springer. pp. 149–168.
Hauser, F., Cazzamali, G., Williamson, M., Blenau, W. and Grimmelikhuijzen, C.J. 2006. A review of neurohormone GPCRs present in the fruitfly Drosophila melanogaster and the honey bee Apis mellifera. Progress in Neurobiology, 80:1–19. https://doi.org/10.1016/j.pneurobio.2006.07.005
Helinski, M.E.H., Deewatthanawong, P., Sirot, L.K., Wolfner, M.F. and Harrington, L.C. 2012. Duration and dose-dependency of female sexual receptivity responses to seminal fuid proteins in Aedes albopictus and Ae. aegypti mosquitoes. Journal of Insect Physiology, 58: 1307–1313.
Hitzemann, R., Bottomly, D., Darakjian, P., Walter, N., Iancu, O., Searles, R., et al. 2013. Genes, behavior and next-generation RNA sequencing. Genes, Brain and Behavior, 12(1):1–12.
Huang, D.W., Sherman, B.T. and Lempicki, R. 2009a. Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nature Protocols, 4: 44–57.
Huang, D.W., Sherman, B.T. and Lempicki, R.A. 2009b. Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic Acids Research, 37: 1–13.
Jasper, W.C., Brutscher, L.M., Grozinger, C.M. and Niño, E.L. 2020. Injection of seminal fluid into the hemocoel of honey bee queens (Apis mellifera) can stimulate post-mating changes. Scientific Reports, 10: 11990.
K-I, H., Sasaki, K. and Nagao, T. 2005 Depression of brain dopamine and its metabolite after mating in European honeybee (Apis mellifera) queens. Naturwissenschaften, 92(7): 310–3. https://doi.org/10.1007/s00114-005-0631-3
Kocher, S., Tarpy, D. and Grozinger, C. 2010. The effects of mating and instrumental insemination on queen honey bee flight behaviour and gene expression. Insect Molecular Biology, 19(2): 153–62
Koywiwattrakul, P., Thompson, G.J., Sitthipraneed, S., Oldroyd, B.P. and Maleszka, R. 2005. Effects of carbon dioxide narcosis on ovary activation and gene expression in worker honeybees, Apis mellifera. Journal of Insect Science, 5.
Love, M.I., Huber, W. and Anders, S. 2014. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biology, 15(12): 550.
Macedo, L.M.F., Nunes, F.M.F., Freitas, F.C.P., Pires, C.V., Tanaka, E.D., Martins, J.R., Piulachs, M.D., Cristino, A.S., Pinheiro, D.G. and Simões, Z.L.P. 2016. MicroRNA signatures characterizing caste-independent ovarian activity in queen and worker honeybees (Apis mellifera L.). Insect Molecular Biology, 25: 216–226.
Manfredini, F., Brown, M.J.F., Vergoz, V. and Oldroyd, B.P. 2015. RNA-sequencing elucidates the regulation of behavioural transitions associated with the mating process in honey bee queens. BMC Genomics, 16: 563. https://doi.org/10.1186/s12864-015-1750-7
Niño, E.L., Tarpy, D.R. and Grozinger, C. 2011. Genome-wide analysis of brain transcriptional changes in honey bee (Apis mellifera L.) queens exposed to. Insect Molecular Biology, 20: 387–398.
Niño, E.L., Tarpy, D.R. and Grozinger, C.M. 2013. Diferential efects of insemination volume and substance on reproductive changes in honey bee queens (Apis mellifera L.). Insect Molecular Biology, 22: 233–244.
Pan, Q., Shai, O., Lee, L.J., Frey, B.J. and Blencowe, B.J. 2008. Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing. Nature Genetics, 40(12): 1413–5.
Pflugfelder, J. and Koeniger, N. 2003. Fight between virgin queens (Apis mellifera) is initiated by contact with the dorsal abdominal surface. Apidologie, 34(3): 249–56.
Richard, F.J., Tarpy, D.R. and Grozinger, C.M. 2007. Effects of insemination quantity on honey bee queen physiology. PLoS ONE, 2(10): e980.
Saito, R., Smoot, M. E., Ono, K., Ruscheinski, J., Wang, P. L., Lotia, S., et al. (2012) A travel guide to Cytoscape plugins. Nature Methods, 9: 1069–1076. https://doi.org/10.1038/nmeth.2212
Sammeth, M., Foissac, S. and Guigo, R. (2008) A general definition and nomenclature for alternative splicing events. PLoS Computational Biology, 4(8): e1000147. https://doi.org/10.1371/journal.pcbi.1000147
Smoot, M.E., Ono, K., Ruscheinski, J., Wang, P.L. and Ideker, T. 2011. Cytoscape 2.8: new features for data integration and network visualization. Bioinformatics, 27: 431–432. https://doi.org/10.1093/bioinformatics/btq675
Tahmasebi, Gh. and Porgharaei, H. 2000. Investigating the role of bee in pollination and increasing the production of agricultural products in Iran. Agricultural Economics and Development, 8(30):131–144.
Tarpy, D.R. and Nielsen, D.I. 2002. Sampling error, effective paternity and estimating the genetic structure of honey bee colonies (Hymenoptera: Apidae). Annals of the Entomological Society of America, 95: 513–528.
Vergoz, V., Lim, J. and Oldroyd, B. 2012. Biogenic amine receptor gene expression in the ovarian tissue of the honey bee Apis mellifera. Insect Molecular Biology, 21(1): 21–9.
Vergoz, V., Lim, J., Duncan, M., Cabanes, G. and Oldroyd, B.P. 2012 Effects of natural mating and CO2 narcosis on biogenic amine receptor gene expression in the ovaries and brain of queen honey bees, Apis mellifera. Insect Molecular Biology, 21(6): 558–67.
Szklarczyk D, Franceschini A, Wyder S, Forslund K, Heller D, Huerta-Cepas J, et al. 2015. STRING v10: protein-protein interaction networks, integrated over the tree of life. Nucleic Acids Research, 43: 447–452.
Journal of Agricultural Science and Technology
Volume 25, Issue 4
May and June 2023
Pages 863-876