Development of Teleost Intermuscular Bones Undergoing Intramembranous Ossification Based on Histological-Transcriptomic-Proteomic Data
Abstract
:1. Introduction
2. Results
2.1. Histological Structures
2.2. General Proteome Analysis During IB Development
2.3. Comparative Analysis of Differentially Expressed Proteins in Different Comparison Groups
2.4. Correlation Analysis of Protein and mRNA
2.5. Network Analysis and Functional Annotation of Altered Bone-Regulated Proteins During IB Development
2.6. Expression Validation of Bone-Related Genes
2.7. Pathway Inhibitor Analysis
3. Discussion
4. Methods
4.1. Methods
4.2. Fish Euthanasia
4.3. Histological Analysis
4.4. iTRAQ-Based Proteomics Analysis
4.4.1. Protein Extraction and iTRAQ Labeling
4.4.2. SCX Fractionation and LC-MS/MS Analysis
4.5. Protein Identification and Proteome Analyses
4.6. Enrichment Analysis
4.7. Integrative Analysis of Proteomic and Transcriptomic Data
4.8. Validation of Differentially Expressed Genes
4.9. Pathway Inhibitor Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
Abbreviations
DMSO | diluted in dimethyl sulfoxide; |
dph | days post-hatch; |
GO | gene ontology; |
HE | hematoxylin-eosin; |
IB | intermuscular bone; |
ICAT | isotope-code affinity tags; |
iTRAQ | Isobaric Tag for Relative Absolute Quantitation; |
KEGG | Kyoto encyclopedia of genes and genomes; |
mRNA | messenger RNA; |
MS-222 | Tricaine methanesulfonate; |
qRT-PCR | Quantitative real-time polymerase chain reaction; |
SILAC | stable isotopes labeling by amino acids in cell culture; |
ARS | alizarin red S; |
LC-MS/MS | liquid chromatography-tandem mass spectrometry. |
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NCBI nr Description | Abbreviation | S2-vs-S1 | S3-vs-S2 | S4-vs-S3 | |||
---|---|---|---|---|---|---|---|
Protein | mRNA | Protein | mRNA | Protein | mRNA | ||
Calsequestrin 1a precursor | casq1a | 0.69 | 0.22 | 0.06 | −0.13 | −0.15 | 0.47 |
Calcium/calmodulin-dependent protein kinase type II subunit beta-like | camk | 0.93 | 0.93 | −0.17 | 0.77 | 0.30 | −1.28 |
Parvalbumin | pvalb | 0.68 | 1.27 | 1.32 | 0.73 | 0.11 | 1.66 |
Calreticulin precursor | calrl | −0.38 | −1.71 | −0.03 | −4.08 | −0.12 | 1.03 |
Annexin A1 | anxa1 | −0.12 | 0.09 | 0.21 | −0.05 | −0.43 | 0.43 |
Annexin A2a | anxa2a | −0.49 | 0.17 | 0.11 | −0.13 | −0.15 | 0.75 |
Annexin A5 | anxa5 | −0.40 | −0.22 | 0.54 | −0.88 | 0.08 | −0.04 |
Ectonucleoside triphosphate diphosphohydrolase 5-like | entpd5 | 0.18 | 0.67 | 0.19 | −0.18 | −0.04 | −0.23 |
Beta-catenin-like protein 1 | β-catenin | −1.09 | −0.49 | −0.07 | −0.42 | 0.11 | −0.29 |
Transforming growth factor, beta-induced | TGF-β | −0.71 | −0.02 | −0.04 | −1.25 | −0.09 | 0.47 |
Tenascin C | tnc | −0.40 | −1.08 | −0.09 | −1.48 | 0.06 | 0.25 |
Signal transducer and activator of transcription 1-alpha/beta | stat1 | −0.97 | 0.24 | −0.03 | 0.72 | 0.28 | −1.12 |
Collagen alpha-1(IX) chain-like, partial | col9α1 | −3.32 | −9.38 | 0.73 | 0.00 | −0.56 | 0.00 |
Laminin subunit beta-2 precursor | lamb2 | −0.07 | −0.42 | 0.01 | −0.002 | −0.64 | −0.31 |
nidogen-1-like | nid1 | 0.11 | −0.58 | 0.04 | −0.67 | 0.15 | −0.45 |
Collagen alpha-3(VI) chain-like | col6α3 | 0.57 | 1.67 | 0.56 | 0.06 | −0.18 | −1.12 |
Matrix metalloproteinase-14 precursor | mmp14b | 0.44 | −0.04 | −0.17 | −1.12 | 0.11 | −0.40 |
Decorin variant 1 | dcn | 0.04 | 0.32 | 0.37 | −1.60 | −0.06 | 0.63 |
Cathepsin K | ctsk | −0.94 | −0.22 | 0.82 | −0.53 | −0.79 | −0.62 |
Prostaglandin E synthase 2-like | PGE2 | −0.22 | −0.40 | 0.12 | −0.13 | −0.20 | −0.38 |
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Nie, C.-H.; Wan, S.-M.; Liu, Y.-L.; Liu, H.; Wang, W.-M.; Gao, Z.-X. Development of Teleost Intermuscular Bones Undergoing Intramembranous Ossification Based on Histological-Transcriptomic-Proteomic Data. Int. J. Mol. Sci. 2019, 20, 4698. https://doi.org/10.3390/ijms20194698
Nie C-H, Wan S-M, Liu Y-L, Liu H, Wang W-M, Gao Z-X. Development of Teleost Intermuscular Bones Undergoing Intramembranous Ossification Based on Histological-Transcriptomic-Proteomic Data. International Journal of Molecular Sciences. 2019; 20(19):4698. https://doi.org/10.3390/ijms20194698
Chicago/Turabian StyleNie, Chun-Hong, Shi-Ming Wan, Yu-Long Liu, Han Liu, Wei-Min Wang, and Ze-Xia Gao. 2019. "Development of Teleost Intermuscular Bones Undergoing Intramembranous Ossification Based on Histological-Transcriptomic-Proteomic Data" International Journal of Molecular Sciences 20, no. 19: 4698. https://doi.org/10.3390/ijms20194698
APA StyleNie, C. -H., Wan, S. -M., Liu, Y. -L., Liu, H., Wang, W. -M., & Gao, Z. -X. (2019). Development of Teleost Intermuscular Bones Undergoing Intramembranous Ossification Based on Histological-Transcriptomic-Proteomic Data. International Journal of Molecular Sciences, 20(19), 4698. https://doi.org/10.3390/ijms20194698