The manuscript mentions that similar studies have been performed in ventricles. Discuss how the results compare.

Throughout the authors use the term ‘expression’ to describe protein abundance, and at times mention ‘upregulation’. Expression can refer to a snap-shot of abundance, hence abundance would be a more accurate terminology. When referencing studies, whether there is a change in mRNA or protein should be clarified.

The NaCl extract was noted to likely contain newly synthesised proteins – hence in this context discuss synthesis versus degradation and turnover (i.e kinetics) in comparison to static ‘abundance’ values and how this could be addressed in future studies.

Regarding the discussion on PLOD3 – if this is increased with age it would be the newly synthesised collagens that have increased hydroxylation and therefore more capacity to be crosslinked – but these may be in the soluble fraction until they are crosslinked (or may be turned over on a circadian basis) and there is no information on the dynamics and kinetics of this process. This should be clarified.

The authors note a decrease in collagens with ageing – could this decrease be due to decreased extractablity/solubility due to increased crosslinking in aged tissues? Were tissues completed solubilised and/or did dry weight change?

The decrease in collagens with age has been noted in C.Elegans (suggest citing work from the Ewald group).

Quantification of N-glycosylation sites is mentioned in the abstract but the relevant figure is in supplementary data. Please harmonise by removing text from abstract or including supplementary in main figures.

Page 3, Line 24 – AF is not ‘another example’ as it was already mentioned in line 10.