Transcriptome Analysis Reveals the Gene Expression Changes in the Silkworm (Bombyx mori) in Response to Hydrogen Sulfide Exposure
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Silkworm Rearing and H2S Exposure
2.2. RNA Extraction
2.3. Sequencing, Data Processing, and Assembly
2.4. Functional Annotation and Enrichment Analysis of DEGs
2.5. Quantitative Real-Time PCR (qRT-PCR)
2.6. Statistical Analysis
3. Results
3.1. Transcriptome Sequencing and Assembly
3.2. DEG Identification and Analysis
3.3. GO Analysis of DEGs
3.4. KEGG Pathway Analysis
3.5. The Validation of DEGs by qRT-PCR
3.6. The Expression Pattern Analysis of DEGs
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
- Goldsmith, M.R.; Shimada, T.; Abe, H. The genetics and genomics of the silkworm, Bombyx mori. Annu. Rev. Entomol. 2005, 50, 71–100. [Google Scholar] [CrossRef] [PubMed]
- Liu, L.; Zhang, P.; Gao, Q.; Feng, X.; Han, L.; Zhang, F.; Bai, Y.; Han, M.; Hu, H.; Dai, F.; et al. Comparative transcriptome analysis reveals bmo-miR-6497-3p regulate circadian clock genes during the embryonic diapause induction process in bivoltine silkworm. Insects 2021, 12, 739. [Google Scholar] [CrossRef] [PubMed]
- Zhu, Z.; Tan, Y.; Xiao, S.; Guan, Z.; Zhao, W.; Dai, Z.; Liu, G.; Zhang, Z. Solitary living brings a decreased weight and an increased agility to the domestic silkworm, Bombyx mori. Insects 2021, 12, 809. [Google Scholar] [CrossRef] [PubMed]
- Lu, Z.Y.; Meng, Z.; Wen, M.Y.; Kang, X.; Zhang, Y.; Liu, Q.; Zhao, P.; Xia, Q. Overexpression of BmFoxO inhibited larval growth and promoted glucose synthesis and lipolysis in silkworm. Mol. Genet. Genomics. 2019, 294, 1375–1383. [Google Scholar] [CrossRef] [PubMed]
- Zheng, S.F.; Jin, X.; Chen, M.H.; Shi, Q.; Zhang, H.; Xu, S. Hydrogen sulfide exposure induces jejunum injury via CYP450s/ROS pathway in broilers. Chemosphere 2019, 214, 25–34. [Google Scholar] [CrossRef]
- Magierowski, M.; Magierowska, K.; Hubalewska-Mazgaj, M.; Surmiak, M.; Sliwowski, Z.; Wierdak, M.; Kwiecien, S.; Chmura, A.; Brzozowski, T. Cross-talk between hydrogen sulfide and carbon monoxide in the mechanism of experimental gastric ulcers healing, regulation of gastric blood flow and accompanying inflammation. Biochem. Pharmacol. 2018, 149, 131–142. [Google Scholar] [CrossRef] [PubMed]
- Miller, D.L.; Roth, M.B. Hydrogen sulfide increases thermotolerance and lifespan in Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA 2007, 104, 20618–20622. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Budde, M.W.; Roth, M.B. Hydrogen sulfide increases hypoxia-inducible factor-1 activity independently of von Hippel-Lindau tumor suppressor-1 in C. elegans. Mol. Biol. Cell 2010, 21, 212–217. [Google Scholar] [CrossRef] [Green Version]
- Qabazard, B.; Li, L.; Gruber, J.; Peh, M.T.; Ng, L.F.; Kumar, S.D.; Rose, P.; Tan, C.H.; Dymock, B.W.; Wei, F.; et al. Hydrogen sulfide is an endogenous regulator of aging in Caenorhabditis elegans. Antioxid. Redox. Signal. 2014, 20, 2621–2630. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wei, Y.; Kenyon, C. Roles for ROS and hydrogen sulfide in the longevity response to germline loss in Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA 2016, 113, E2832–E2841. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Kabil, H.; Kabil, O.; Banerjee, R.; Harshman, L.G.; Pletcher, S.D. Increased transsulfuration mediates longevity and dietary restriction in Drosophila. Proc. Natl. Acad. Sci. USA 2011, 108, 16831–16836. [Google Scholar] [CrossRef] [Green Version]
- Zhong, J.F.; Wang, S.P.; Shi, X.X.; Mu, L.L.; Li, G.Q. Hydrogen sulfide exposure increases desiccation tolerance in Drosophila melanogaster. J. Insect Physiol. 2010, 56, 1777–1782. [Google Scholar] [CrossRef]
- Cao, Y.Y.; Peng, L.L.; Jiang, L.; Thakur, K.; Hu, F.; Tang, S.M.; Wei, Z.J. Evaluation of the metabolic effects of hydrogen sulfide on the development of Bombyx mori (Lepidoptera: Bombycidae), using liquid chromatography-mass spectrometry-based metabolomics. J. Insect Sci. 2020, 20, 4. [Google Scholar] [CrossRef]
- Jiang, L.; Peng, L.L.; Cao, Y.Y.; Thakur, K.; Tang, S.M.; Hu, F.; Wei, Z.J. Transcriptome analysis reveals gene expression changes of the fat body of silkworm (Bombyx mori L.) in response to selenium treatment. Chemosphere 2020, 245, 125660. [Google Scholar] [CrossRef]
- Martin, M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet. J. 2011, 17, 10–12. [Google Scholar] [CrossRef]
- Pertea, M.; Kim, D.; Pertea, G.M.; Leek, J.T.; Salzberg, S.L. Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown. Nat. Protoc. 2016, 11, 1650–1667. [Google Scholar] [CrossRef]
- Beekman, R.; Chapaprieta, V.; Russiñol, N.; Vilarrasa-Blasi, R.; Verdaguer-Dot, N.; Martens, J.H.A.; Duran-Ferrer, M.; Kulis, M.; Serra, F.; Javierre, B.M.; et al. The reference epigenome and regulatory chromatin landscape of chronic lymphocytic leukemia. Nat. Med. 2018, 24, 868–880. [Google Scholar] [CrossRef]
- Xia, Q.; Zhou, Z.; Lu, C.; Cheng, D.; Dai, F.; Li, B.; Zhao, P.; Zha, X.; Cheng, T.; Chai, C.; et al. A draft sequence for the genome of the domesticated silkworm (Bombyx mori). Science 2004, 306, 1937–1940. [Google Scholar]
- Kim, D.; Langmead, B.; Salzberg, S.L. HISAT: A fast spliced aligner with low memory requirements. Nat. Methods 2015, 12, 357–360. [Google Scholar] [CrossRef] [Green Version]
- Pertea, M.; Pertea, G.M.; Antonescu, C.M.; Chang, T.C.; Mendell, J.T.; Salzberg, S.L. StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat. Biotechnol. 2015, 33, 290–295. [Google Scholar] [CrossRef] [Green Version]
- Robinson, M.D.; McCarthy, D.J.; Smyth, G.K. edgeR: A Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics. 2010, 26, 139–140. [Google Scholar] [CrossRef] [Green Version]
- Kanehisa, M.; Araki, M.; Goto, S.; Hattori, M.; Hirakawa, M.; Itoh, M.; Katayama, T.; Kawashima, S.; Okuda, S.; Tokimatsu, T.; et al. KEGG for linking genomes to life and the environment. Nucleic Acids Res. 2008, 36, D480–D484. [Google Scholar] [CrossRef]
- Zhang, F.; Zhang, Y.Y.; Ma, R.H.; Thakur, K.; Han, J.; Hu, F.; Zhang, J.G.; Wei, Z.J. Multi-omics reveals the anticancer mechanism of asparagus saponin-asparanin A on endometrial cancer Ishikawa cells. Food Funct. 2021, 12, 614–632. [Google Scholar] [CrossRef]
- Wang, J.J.; Bai, W.W.; Zhou, W.; Liu, J.; Chen, J.; Liu, X.Y.; Xiang, T.T.; Liu, R.H.; Wang, W.H.; Zhang, B.L.; et al. Transcriptomic analysis of two Beauveria bassiana strains grown on cuticle extracts of the silkworm uncovers their different metabolic response at early infection stage. J. Invertebr. Pathol. 2017, 145, 45–54. [Google Scholar] [CrossRef]
- Adam, G.M.; Naama, K. NADH ties One-Carbon metabolism to cellular respiration. Cell Press. 2020, 31, 660–662. [Google Scholar]
- Chen, Q.M.; Ma, Z.G.; Wang, X.; Li, Z.; Zhang, Y.; Ma, S.; Zhao, P.; Xia, Q. Comparative proteomic analysis of silkworm fat body after knocking out fibroin heavy chain gene: A novel insight into cross-talk between tissues. Funct. Integr. Genomics. 2015, 15, 611–637. [Google Scholar] [CrossRef]
- Long, W.; Wu, J.; Shen, G.; Zhang, H.; Liu, H.; Xu, Y.; Gu, J.; Jia, L.; Lin, Y.; Xia, Q. Estrogen-related receptor participates in regulating glycolysis and influences embryonic development in silkworm Bombyx mori. Inesct Mol. Biol. 2020, 29, 160–169. [Google Scholar] [CrossRef]
- Efe, S.; Duvernell, D.D.; Matzkin, M.L.; Duan, Y.H.; Zhu, C.T.; Verrelli, B.C.; Eanes, W.F. Single-Locus latitudinal clines and their relationship to temperate adaptation in metabolic genes and derived alleles in Drosophila melanogaster. Genet. Soc. Am. 2004, 168, 923–931. [Google Scholar]
- Long, L.; Wang, C.C.; Zhao, X.Y.; Guan, J.X.; Lei, C.L.; Huang, Q.Y. Isocitrate dehydrogenase-mediated metabolic disorders disrupt active immunization against fungal pathogens in eusocial termites. J. Pest. Sci. 2020, 93, 291–301. [Google Scholar]
- Meyer, S.; Gessner, D.K.; Wen, C.P.; Most, E.; Liebisch, G.; Zorn, H.; Ringseis, R.; Eder, K. The Antisteatotic and Hypolipidemic Effect of Insect Meal in Obese Zucker Rats is Accompanied by Profound Changes in Hepatic Phospholipid and 1-Carbon Metabolism. Mol. Nutr. Food Res. 2019, 63, e1801305. [Google Scholar] [CrossRef]
- Shi, A.B.; Chen, C.C.; Banerjee, R.; Glodowski, D.; Audhya, A.; Rongo, C.; Grant, B.D. EHBP-1 functions with RAB-10 during endocytic recycling in Caenorhabditis elegans. Mol. Biol. Cell. 2010, 21, 2930–2943. [Google Scholar] [CrossRef] [Green Version]
- Le Roy, C.; Wrana, J.L. Clathrin- and non-clathrin-mediated endocytic regulation of cell signaling. Nat. Rev. Mol. Cell Biol. 2005, 6, 112–126. [Google Scholar] [CrossRef]
- Nichols, B. Caveosomes and endocytosis of lipid rafts. J. Cell Sci. 2003, 116, 4707–4714. [Google Scholar] [CrossRef] [Green Version]
- Stenmark, H. Rab GTPases as coordinators of vesicle traffic. Nat. Rev. Mol. Cell Biol. 2009, 10, 513–525. [Google Scholar] [CrossRef]
- Glodowski, D.R.; Chen, C.C.; Schaefer, H.; Grant, B.D.; Rongo, C. RAB-10 regulates glutamate receptor recycling in a cholesterol-dependent endocytosis pathway. Mol. Biol. Cell. 2007, 18, 4387–4396. [Google Scholar] [CrossRef] [Green Version]
- Tian, J.H.; Xue, B.; Hu, J.H.; Li, J.X.; Cheng, X.Y.; Hu, J.S.; Li, F.C.; Chen, Y.H.; Li, B. Exogenous substances regulate silkworm fat body protein synthesis through MAPK and PI3K/Akt signaling pathways. Chemosphere 2017, 171, 202–207. [Google Scholar] [CrossRef]
- Zanardo, R.C.; Brancaleone, V.; Distrutti, E.; Fiorucci, S.; Cirino, G.; Wallace, J.L. Hydrogen sulfide is an endogenous modulator of leukocyte-mediated inflammation. FASEB J. 2006, 20, 2118–2120. [Google Scholar] [CrossRef]
- Jia, Q.; Wang, T.T.; Wang, X.Y.; Xu, H.; Liu, Y.; Wang, Y.; Shi, Q.; Liang, Q. Astragalin suppresses inflammatory responses and bone destruction in mice with collagen-induced arthritis and in human fibroblast-likesynoviocytes. Front. Pharmacol. 2019, 10, 94. [Google Scholar] [CrossRef] [Green Version]
- Hu, W.; Wang, X.; Ma, S.; Peng, Z.; Cao, Y.; Xia, Q. CRISPR-Mediated endogenous activation of fibroin heavy chain gene triggers cellular stress responses in Bombyx mori Embryonic Cells. Insects 2021, 12, 552. [Google Scholar] [CrossRef]
- Kunz, R.I.; Brancalhao, R.M.; Ribeiro, L.F.; Natali, M.R. Silkworm sericin: Properties and biomedical applications. Biomed. Res. Int. 2016, 2016, 8175701. [Google Scholar] [CrossRef] [Green Version]
- Ni, M.; Zhang, H.; Li, F.C.; Wang, B.B.; Xu, K.Z.; Shen, W.D.; Li, B. Nanoparticulate anatase TiO2 (TiO2NPs) upregulates the expression of silkworm (Bombyx mori) neuropeptide receptor and promotes silkworm feeding, growth, and silking. Peptides 2015, 68, 64–71. [Google Scholar] [CrossRef]
Samples | FB_Control_1 | FB_Control_2 | FB_Control_3 | FB_H2S_1 | FB_H2S_2 | FB_H2S_3 |
---|---|---|---|---|---|---|
Raw reads | 52,598,692 | 49,647,104 | 50,987,488 | 50,804,424 | 53,068,042 | 51,108,946 |
(7.89 Gb) | (7.45 Gb) | (7.65 Gb) | (7.62 Gb) | (7.96 Gb) | (7.67 Gb) | |
High quality reads | 49,443,624 | 49,144,228 | 50,029,926 | 50,357,636 | 52,490,302 | 50,286,178 |
(7.42 Gb) | (7.37 Gb) | (7.50 Gb) | (7.55 Gb) | (7.87 Gb) | (7.54 Gb) | |
High quality reads ratio (%) | 94.00 | 98.99 | 98.12 | 99.12 | 98.91 | 98.39 |
Q30 (%) | 98.53 | 99.08 | 99.07 | 98.93 | 98.98 | 99.12 |
GC content (%) | 50 | 48 | 49 | 48 | 48.50 | 48 |
Total mapped | 46,077,049 | 47,843,354 | 48,788,614 | 49,023,730 | 51,140,817 | 49,141,008 |
Pathway ID | Pathway Name | Number of DEGs | p-Value |
---|---|---|---|
ko03010 | Ribosome | 33 | <0.01 |
ko04510 | Focal adhesion | 20 | <0.01 |
ko05132 | Salmonella infection | 7 | <0.01 |
ko04370 | VEGF signaling pathway | 6 | <0.01 |
ko04810 | Regulation of actin cytoskeleton | 15 | <0.01 |
ko00360 | Phenylalanine metabolism | 7 | <0.01 |
ko04530 | Tight junction | 13 | <0.01 |
ko04668 | TNF signaling pathway | 4 | 0.01 |
ko00010 | Glycolysis/Gluconeogenesis | 11 | 0.01 |
ko04144 | Endocytosis | 35 | 0.01 |
ko00562 | Inositol phosphate metabolism | 12 | 0.01 |
ko00130 | Ubiquinone and other terpenoid-quinone biosynthesis | 6 | 0.01 |
ko04380 | Osteoclast differentiation | 3 | 0.01 |
ko04670 | Leukocyte transendothelial migration | 3 | 0.01 |
ko00410 | beta-Alanine metabolism | 7 | 0.01 |
ko05100 | Bacterial invasion of epithelial cells | 4 | 0.02 |
ko04611 | Platelet activation | 3 | 0.02 |
ko04520 | Adherens junction | 7 | 0.02 |
ko04621 | NOD-like receptor signaling pathway | 4 | 0.02 |
ko04391 | Hippo signaling pathway-fly | 19 | 0.02 |
ko04010 | MAPK signaling pathway | 16 | 0.03 |
ko00340 | Histidine metabolism | 4 | 0.04 |
ko05410 | Hypertrophic cardiomyopathy (HCM) | 6 | 0.04 |
ko04064 | NF-kappa B signaling pathway | 2 | 0.04 |
ko00020 | Citrate cycle (TCA cycle) | 9 | 0.04 |
ko04962 | Vasopressin-regulated water reabsorption | 4 | 0.04 |
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Zhang, R.; Cao, Y.-Y.; Du, J.; Thakur, K.; Tang, S.-M.; Hu, F.; Wei, Z.-J. Transcriptome Analysis Reveals the Gene Expression Changes in the Silkworm (Bombyx mori) in Response to Hydrogen Sulfide Exposure. Insects 2021, 12, 1110. https://doi.org/10.3390/insects12121110
Zhang R, Cao Y-Y, Du J, Thakur K, Tang S-M, Hu F, Wei Z-J. Transcriptome Analysis Reveals the Gene Expression Changes in the Silkworm (Bombyx mori) in Response to Hydrogen Sulfide Exposure. Insects. 2021; 12(12):1110. https://doi.org/10.3390/insects12121110
Chicago/Turabian StyleZhang, Rui, Yu-Yao Cao, Juan Du, Kiran Thakur, Shun-Ming Tang, Fei Hu, and Zhao-Jun Wei. 2021. "Transcriptome Analysis Reveals the Gene Expression Changes in the Silkworm (Bombyx mori) in Response to Hydrogen Sulfide Exposure" Insects 12, no. 12: 1110. https://doi.org/10.3390/insects12121110