Background β-catenin is a key component of Wnt signalling pathway which controls brain development and neurogenesis of the adult brain. Within a cell, β-catenin is continuously synthesised and its level is strictly regulated by phosphorylation and subsequent degradation via ubiquitin-proteasome system. Recently, the accumulation of β-catenin has been associated with Huntington disease as a consequence of the impaired degradation caused by mutated huntingtin.
Aims Our study is aimed at monitoring β-catenin level in minipigs transgenic for the N-terminal part of human mutated huntingtin, the model developed in our institute.
Methods Using Western blot and specific antibodies, transgenic animals and their siblings of the same age and the identical genetic background were examined for the expression of β-catenin. Furthermore, phosphorylation of β-catenin was monitored applying the method of phosphoprotein isolation. Finally, β-catenin was isolated by immunoprecipitation and subjected to mass-spectrometric analysis.
Results Two forms of β-catenin with minimal variance in molecular weight were detected in all samples from transgenic and wild-type pigs. Although there were no significant differences in β-catenin of the lower molecular weight, a noticeable decrease of heavier form was observed in transgenic animals. More interestingly, β-catenin of lower molecular weight was substantially phosphorylated suggesting that such modification is not responsible for the presence of heavier β-catenin in control animals. In order to reveal the differences among two observed forms of β-catenin, immunoprecipitation followed by mass spectrometry was performed and results will be discussed.
Conclusion Our results demonstrate the presence of two distinct forms of β-catenin and indicate that a specific form of the protein is almost absent in transgenic animals. We hope that in depth investigation of this protein will extend our knowledge about the pathophysiology of Huntington disease.
- transgenic pig
- mutated huntingtin
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