Combining Multiple Omics with Molecular Dynamics Reveals SCP2-Mediated Cytotoxicity Effects of Aflatoxin B1 in SW480 Cells
Abstract
:1. Introduction
2. Result
2.1. Cell Viability
2.2. Transcriptomic Analysis Based on Differentially Expressed Genes (DEGs)
2.3. Proteomics Analysis Based on Differentially Expressed Proteins (DEPs)
2.4. Analysis of the Metabolomics Data between the Group Treated with AFB1 and the Control Group
2.5. The Potential Target for AFB1 Toxicity
2.6. Oxidative Stress Induced by AFB1
2.7. The Antioxidant Enzyme Activity
2.8. The Exposure to AFB1 Leads to Excessive Accumulation of Intracellular Ca2+
2.9. Fatty Acid Localization
2.10. AFB1-Induced Mitochondrial Dysfunction
2.11. mRNA and Protein Expression Involved in Oxidative Stress and Lipid Signaling Pathways
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Chemical Reagent and Cell Culture
5.2. Cell Viability Assay
5.3. RNA Extraction and Transcriptomics
5.4. Tandem Mass Tag Proteomics Analysis
5.5. Sample Preparation and Analysis for Metabolomics
5.6. Molecular Docking
5.7. Molecular Dynamics (MD)
5.8. ROS Measurement by DCFH-DA
5.9. Assay of Catalase (CAT) and Glutathione S-Transferase (GST)
5.10. Measurement of Cellular Calcium
5.11. Fatty Acid Green Fluorescence Probe
5.12. Detection of Mitochondrial Membrane Potential (MMP)
5.13. Detection of Mitochondrial Permeability Transition Pore (mPTP)-Opening
5.14. Assessment of Mitochondrial Activity
5.15. RT-qPCR
5.16. Western Blotting
5.17. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Protein | PDB ID | Total Score | Crash | Polar | H-Bond Number | Residues Involved in H-Bond Formation | Hydrophobic Contact Number | Residues Involved in Hydrophobic Contacts |
---|---|---|---|---|---|---|---|---|
CAT | 8HID | 7.7967 | −0.2167 | 0 | 3 | Arg72, Arg112 (2 H-bonds) | 10 | Val73, Tyr358, Val74, Arg354, Val146, His75, Phe132, Gly131, Ser114, Ala133 |
SEC61A1 | 8DNV | 5.6242 | −0.7089 | 2.9825 | 0 | − | 7 | Ile292, Ser287, Gln456, Ser180, Gly176, Ser177, Asn288 |
ACSL1 | 2JFK | 5.5354 | −0.9486 | 1.9938 | 1 | Ser581 | 12 | Met620, His683, Val743, Met499, Thr650, Val675, Ala622, Leu767, His580, Ser497, Leu766, Gly498 |
SCP2 | 1IKT | 5.1034 | −0.5708 | 1.8757 | 2 | So4122, Lys115, | 7 | Leu114, Val82, Gln111, Val83, Phe79, Ile15, Val11 |
LPIN1 | 5WGB | 4.8557 | −0.6691 | 3.1075 | 2 | Tyr26, Arg34 | 5 | Val33, Thr415, Glu417, Thr30, Glu416 |
NQO1 | 5EA2 | 4.24 | −0.6254 | 1.0619 | 2 | Lys113, Tyr126 | 4 | Tyr67, Phe120, Glu117, Phe178 |
Gene | Forward Primer (5′-3′) | Reverse Primer (5′-3′) |
---|---|---|
GRP78 | CAGTTGTTACTGTACCAGCCTA (F) | CATTTAGGCCAGCAATAGTTCC (R) |
PERK | CCAGTTTTGTACTCCAATTGCA (F) | CAGATACAGCTGGCCTCTATAC (R) |
IRE1α | CGTGAGCGACAGAATAGAAAAG (F) | GCTTCTTATTTCTCATGGCTCG (R) |
XBP1 | CTTGTAGTTGAGAACCAGGAGT (F) | CCCAACAGGATATCAGACTCTG (R) |
SEC61A1 | AATTTGAAGGTGCTATCATCGC (F) | ACGTATAGAAGAGCTTGATGGG (R) |
CAT | GAGCACAGCATCCAATATTCTG (F) | CTCATTCAGCACGTTCACATAG (R) |
GPX1 | GTTGCCTGGAACTTTGAGAAG (F) | CTCGATGTCAATGGTCTGGAAGG (R) |
GSS | CTCTTTGACATCCACAAGCAAG (F) | AACATGTAGTCTGAGCGATTCA (R) |
NQO1 | CAACCACGAGCCCAGCCAATC (F) | ACTCCACCACCTCCCATCCTTTC (R) |
COX2 | TGTCAAAACCGAGGTGTATGTA (F) | AACGTTCCAAAATCCCTTGAAG (R) |
HO-1 | CCTCCCTGTACCACATCTATGT (F) | GCTCTTCTGGGAAGTAGACAG (R) |
ACSBG1 | AAACAAGATGGCCAATGTGTAC (F) | CGTGATATTGTCTTGACTCAGC (R) |
ACSL5 | ATATGATGCTGAGAACCTAGGC (F) | CATGCTCCACACATTTGAGAAA (R) |
CYP4F3 | AGAAGAAGGGGAGAGGAGGTTGTG (F) | GAGGCGGCAGCAGTTGTCATAG (R) |
CYP4F12 | AAGGCATTGTCAGATGAGGATA (F) | TTCAATCTCTTTAGGATCGCGG (R) |
LPIN1 | GAGCAGCAGAACTCTTCCTAAT (F) | CTTTTGCAATCTACCAGGCTAC (R) |
LDLR | CTGTAGGGGTCTTTACGTGTTC (F) | GTTTTCCTCGTCAGATTTGTCC (R) |
ACSL1 | CTGTGTCATGGAGCTAAAATCG (F) | GTGTTTGCTTGTCCGAAAATTC (R) |
SCP2 | GGAGCTGAGAATTCAAGAGACT (F) | AGTCACCAAAAACATAGCCAAC (R) |
β-ACTIN | GCACTCTTCCAGCCTTCCTTCC (F) | GCGGATGTCCACGTCACACTTC (R) |
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Chen, M.; Wen, J.; Qiu, Y.; Gao, X.; Zhang, J.; Lin, Y.; Wu, Z.; Lin, X.; Zhu, A. Combining Multiple Omics with Molecular Dynamics Reveals SCP2-Mediated Cytotoxicity Effects of Aflatoxin B1 in SW480 Cells. Toxins 2024, 16, 375. https://doi.org/10.3390/toxins16090375
Chen M, Wen J, Qiu Y, Gao X, Zhang J, Lin Y, Wu Z, Lin X, Zhu A. Combining Multiple Omics with Molecular Dynamics Reveals SCP2-Mediated Cytotoxicity Effects of Aflatoxin B1 in SW480 Cells. Toxins. 2024; 16(9):375. https://doi.org/10.3390/toxins16090375
Chicago/Turabian StyleChen, Mengting, Jiaxin Wen, Yiyan Qiu, Xinyue Gao, Jian Zhang, Yifan Lin, Zekai Wu, Xiaohuang Lin, and An Zhu. 2024. "Combining Multiple Omics with Molecular Dynamics Reveals SCP2-Mediated Cytotoxicity Effects of Aflatoxin B1 in SW480 Cells" Toxins 16, no. 9: 375. https://doi.org/10.3390/toxins16090375