Molecular Basis of Unequal Alternative Splicing of Human SCD5 and Its Alteration by Natural Genetic Variations
Abstract
:1. Introduction
2. Results
2.1. Confirmation of Transcript Variants A and B of the SCD5 Gene
2.2. Expression and Distribution of SCD1, SCD5A and SCD5B mRNAs in Human Tissues
2.3. In Silico Analysis of Splice Sites in the Alternative Terminator Region of SCD5
2.4. Minigene Assay of Splice Sites in the Alternative Terminator Region of SCD5
2.5. Effect of Natural Genetic Variants of SCD5A- and SCD5B-Specific Donor and Acceptor Sites on Alternative Splicing
2.5.1. In Silico Analysis
2.5.2. In Vitro Analysis
3. Discussion
4. Materials and Methods
4.1. Chemicals and Materials
4.2. Web-Based Tools for In Silico Analysis
4.3. Expression Plasmid Construction and Mutagenesis
4.4. Cell Culture and Transfection
4.5. Preparation of Cell Lysates
4.6. Immunoblot Analysis
4.7. RNA Isolation, cDNA Synthesis and RT-PCR
4.8. qPCR Analysis
4.9. Statistics
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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SNV ID | Wt | Mut | Affected Exons | Change in | Predicted Consequence in | Impact | ||
---|---|---|---|---|---|---|---|---|
Allele | Amino Acid | Codon | Protein | mRNA | ||||
rs145164872 | A | G | 3/4B | Asn/Asp | Aat/Gat | missense | srv | moderate |
3/4A | Lys/Glu | Aag/Gag | moderate | |||||
rs1430176385 | G | A | 3/4B | – | – | – | sav | high |
rs140750150 | C | T | 3/4B | Thr/Ile | aCa/aTa | missense | srv | moderate |
G | Thr/Arg | aCa/aGa | missense | srv | moderate | |||
rs1250613148 | C | A | 3/4A | – | – | – | spYv, srv | low |
rs1225904796 | C | T | 3/4A | – | – | – | spYv, srv | low |
rs1011850309 | G | C | 3/4A | Lys/Asn | aaG/aaC | missense | srv | moderate |
A | Lys | aaG/aaA | synonymous | srv | low |
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Orosz, G.; Szabó, L.; Bereti, S.; Zámbó, V.; Csala, M.; Kereszturi, É. Molecular Basis of Unequal Alternative Splicing of Human SCD5 and Its Alteration by Natural Genetic Variations. Int. J. Mol. Sci. 2023, 24, 6517. https://doi.org/10.3390/ijms24076517
Orosz G, Szabó L, Bereti S, Zámbó V, Csala M, Kereszturi É. Molecular Basis of Unequal Alternative Splicing of Human SCD5 and Its Alteration by Natural Genetic Variations. International Journal of Molecular Sciences. 2023; 24(7):6517. https://doi.org/10.3390/ijms24076517
Chicago/Turabian StyleOrosz, Gabriella, Luca Szabó, Szanna Bereti, Veronika Zámbó, Miklós Csala, and Éva Kereszturi. 2023. "Molecular Basis of Unequal Alternative Splicing of Human SCD5 and Its Alteration by Natural Genetic Variations" International Journal of Molecular Sciences 24, no. 7: 6517. https://doi.org/10.3390/ijms24076517