Presynaptic signalling in Huntington’s disease

Huntington’s disease (HD) is a heritable neurodegenerative disorder caused by a variable repeat expansion of the CAG codon in exon1 of the huntingtin (Htt) gene. This monogenic mutation results in the expression of mutant (m)Htt protein and causes neurodegeneration, principally in the medium spiny neurons of the striatum. The symptoms of HD include loss of motor coordination (chorea) and cognitive deficits such as loss of memory and concentration. Currently, there are no treatments available that can cure or prevent HD. More research is needed to better understand HD to develop better treatments. As in many neurodegenerative disorders, synaptic dysfunction is a critical early event in the progression of HD. It is known that both protein and mRNA expression of brain derived neurotrophic factor (BDNF) are defective in HD patients and preclinical models. Crucially, members of BDNF signalling pathway regulate a form of presynaptic function called activity dependent endocytosis (ADBE) which is the dominant mode of endocytosis at the presynapse during high frequency stimulation. Previous work in this laboratory showed that ADBE is increased in primary cultured hippocampal and striatal neurons of the HttQ140 knock-in mouse model of HD. Thus the BDNF signalling pathway may link presynaptic dysfunction and HD. Presynaptic function was therefore investigated in primary neuronal cultures and synaptosome preparations from the HttQ140 knock-in mouse model. Using western blotting, there were no detected changes in phosphorylation levels of the downstream BDNF signalling molecules dynamin1 (Dyn1) and glycogen synthase kinase 3 (GSK3) in HttQ140/+ hippocampal neurons compared to Htt+/+. Thus the increase in ADBE exhibited by HttQ140 neurons is not due to the dysregulation of phospho-Dyn1 or phospho-GSK3. Akt1, which is known to be dysregulated in HD, is another downstream member of the BDNF signalling pathway involved in regulating ADBE. Previous work showed that Akt1, which phosphorylates Htt at serine residue 421 (Htt S421), exhibits decreased expression in bulk endosome isolations in HttQ140/Q140 mice. Therefore, ADBE was assayed using tetramethylrhodamine dextran uptake and fluorescence microscopy to determine whether Akt1 function is responsible for impaired ADBE in HttQ140/+ neurons. Reinforcing what has been shown previously, hippocampal HttQ140/+ neurons exhibited increased ADBE compared to Htt+/+ neurons. Interestingly, over-expression of the phospho-mimetic Htt serine 421 aspartate (Htt S421D) was able to rescue the increase in ADBE in HttQ140/+ neurons whilst Htt+/+ neurons were unaffected. Therefore modulation of Htt S421, which is regulated by Akt1, can rescue ADBE in HD neurons. Subsequently, immunoprecipitation experiments were performed using synaptosome preparations from Htt+/+ and HttQ140/+ mice to identify synaptic Htt and phospho-Htt interactors using label-free mass spectroscopy. Protein-protein interaction network and gene ontology analysis were performed to interrogate the potential effect of phospho-dependent alterations in the interactions of Htt and mHtt. This identified target molecules for further research into the mechanisms underlying ADBE dysfunction in HD with the potential to generate novel targets for new therapeutic interventions.

Grantor dc:publisher

The University of Edinburgh

Year dc:date.issued

2025

Author dc:creator

• Runciman, Hamish

Advisors dc:contributor.advisor

• Smillie, Karen
• Cousin, Michael
• Murray, Lyndsay

• Huntington’s disease (HD)
• Presynaptic function
• BDNF signalling pathway
• Activity dependent endocytosis (ADBE)
• Akt1

Language dc:language.iso

en

Identifier URI

http://dx.doi.org/10.7488/era/6044

OAI identifier oai:identifier

oai:era.ed.ac.uk:1842/43508