Loss of SPRED3 Causes Primary Hypothyroidism and Alters Thyroidal Expression of Autophagy Regulators LC3, p62, and ATG5 in Mice
Abstract
1. Introduction
2. Results
2.1. Generation and Validation of SPRED3 Knockout Mice
- A reporter trap element including:
- ○
- A splice acceptor (SA) sequence from the mouse engrailed-2 gene.
- ○
- A lacZ reporter gene for β-galactosidase expression.
- ○
- A polyadenylation (Poly (A)) signal derived from Simian Virus 40, which truncates the endogenous transcription.
- A neomycin resistance gene (neo) driven by a phosphoglycerate kinase (PGK) promoter, flanked by FRT sites for conditional removal.
- Three loxP sites enabling the potential for conditional gene deletion [17].
2.2. SPRED3 Deficiency Causes Growth Retardation and Deviations from Mendelian Ratios
2.3. SPRED3 Deficiency Does Not Alter Cardiac Performance
2.4. Scrutiny of Tissue-Specific Spred3 Promoter Activity
2.5. Hormonal Profiling Reveals Primary Hypothyroidism in SPRED3 KO Mice
2.6. ERK Signaling in the Thyroid Gland Is Reduced in SPRED3 KO Mice
2.7. SPRED3 Deficiency Alters Autophagy-Related Protein Expression in the Thyroid Gland
3. Discussion
4. Materials and Methods
4.1. SPRED3 Mutant Mice
4.2. X-Gal Staining
4.3. Genotyping of Mice
4.4. RNA Extraction and RT-qPCR Analyses
4.5. Western Blotting and Densitometric Quantification of Detected Bands
4.6. Examination of Serum Hormone Levels
4.7. Invasive Hemodynamics
4.8. Co-Immunoprecipitation Assays
4.9. ChimeraX
4.10. Statistical Analysis
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Hemodynamic Parameters (Isoflurane Anesthesia, ±SEM) | WT (n = 13) | SPRED3-KO (n = 12) |
---|---|---|
HR (min−1) | 488 ± 12.5 | 497 ± 17.8 |
SV (µL) | 15.3 ± 0.6 | 15.5 ± 0.7 |
ESV (µL) | 17.4 ± 1.2 | 16.5 ± 1.5 |
EDV (µL) | 30.8 ± 1.6 | 30.5 ± 1.3 |
EF (%) | 49.6 ± 2.1 | 51.9 ± 2.9 |
CO (µL min−1) | 7426.6 ± 212.9 | 7807.6 ± 542.2 |
ESP (mmHg) | 95.6 ± 2.7 | 94.8 ± 3.5 |
EDP (mmHg) | 8.5 ± 1.3 | 6.4 ± 0.7 |
SW (mmHg × µL) | 1208.0 ± 63.6 | 1255.3 ± 70.8 |
V@dP/dt max (µL) | 31.3 ± 1.4 | 30.4 ± 1.2 |
V@dP/dt min (µL) | 16.9 ± 1.2 | 16.0 ± 1.3 |
Ea (mmHg µL−1) | 6.3 ± 0.3 | 6.2 ± 0.4 |
Systolic indices | ||
dP/dt max (mmHg s−1) | 7657.1 ± 263.5 | 8465.8 ± 682.4 |
dV/dt max (µL s−1) | 642.9 ± 25.3 | 693.2 ± 37.8 |
Diastolic indices | ||
-dP/dt min (-mmHg s−1) | 8385.2 ± 426.8 | 9284.7 ± 637.0 |
Tau (W) (ms) | 6.7 ± 0.4 | 5.9 ± 0.4 |
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Dogan, C.; Haas, L.; Holzapfel, R.; Schmitt, F.; Hepbasli, D.; Ullrich, M.; Bösl, M.R.; Abeßer, M.; Schuh, K.; Gredy, S. Loss of SPRED3 Causes Primary Hypothyroidism and Alters Thyroidal Expression of Autophagy Regulators LC3, p62, and ATG5 in Mice. Int. J. Mol. Sci. 2025, 26, 7660. https://doi.org/10.3390/ijms26157660
Dogan C, Haas L, Holzapfel R, Schmitt F, Hepbasli D, Ullrich M, Bösl MR, Abeßer M, Schuh K, Gredy S. Loss of SPRED3 Causes Primary Hypothyroidism and Alters Thyroidal Expression of Autophagy Regulators LC3, p62, and ATG5 in Mice. International Journal of Molecular Sciences. 2025; 26(15):7660. https://doi.org/10.3390/ijms26157660
Chicago/Turabian StyleDogan, Celine, Luisa Haas, Rebecca Holzapfel, Franziska Schmitt, Denis Hepbasli, Melanie Ullrich, Michael R. Bösl, Marco Abeßer, Kai Schuh, and Sina Gredy. 2025. "Loss of SPRED3 Causes Primary Hypothyroidism and Alters Thyroidal Expression of Autophagy Regulators LC3, p62, and ATG5 in Mice" International Journal of Molecular Sciences 26, no. 15: 7660. https://doi.org/10.3390/ijms26157660
APA StyleDogan, C., Haas, L., Holzapfel, R., Schmitt, F., Hepbasli, D., Ullrich, M., Bösl, M. R., Abeßer, M., Schuh, K., & Gredy, S. (2025). Loss of SPRED3 Causes Primary Hypothyroidism and Alters Thyroidal Expression of Autophagy Regulators LC3, p62, and ATG5 in Mice. International Journal of Molecular Sciences, 26(15), 7660. https://doi.org/10.3390/ijms26157660