Write here any general comments you might have about the research approach.
This is an extensive study in which the authors use numerous orthogonal methods to investigate HTT protein structure-function relationships and provide insight into consequences of polyglutamine expansion at the molecular level.
HTT phosphorylation status and polyglutamine tract length are known to modulate various cellular functions in many different HD models. However, it remains unclear how single PTM sites and expansion of the polyglutamine tract, distal to the bulk globular protein structure, could mediate the large conformational changes of the HTT molecule detailed in the manuscript. We believe there is insufficient focus on establishing more precise mechanisms by which these changes might be mediated through the protein structure.
Analysis of intermediate polyQ lengths would further strengthen the work i.e. Q36 (at risk) and Q42 (common patient Q-tract length) when relating structure to function. Q78 represents an extreme expansion rarely seen in patients so the physiological relevance of assessing this protein compared to more disease relevant Q-tract expansion lengths is not clear.
It is unclear whether chemically cross-linked HTT is used in all experiments detailed or just in the cryoEM analyses. This should be clearly explained before each experiment as this is a critical detail.