Detection, visualization and quantification of protein complexes in human Alzheimer’s disease brains using proximity ligation assay

Detection, visualization and quantification of protein complexes in human Alzheimer's disease brains using proximity ligation assay

AUTHORS

Romero-Fernandez WWilber , Carvajal-Tapia CCristian , Prusky AAlex , Katdare KAKetaki A , Wang EEmmeline , Shostak AAlena , Ventura-Antunes LLissa , Harmsen HJHannah J , Lippmann ESEthan S , Fuxe KKjell , MacGurn JAJason A , Borroto-Escuela DODasiel O , Schrag MSMatthew S . Scientific reports. 2023 07 24; 13(1). 11948

PMID: 37488165[PubMed].

ABSTRACT

Examination of healthy and diseased human brain is essential to translational neuroscience. Protein-protein interactions play a pivotal role in physiological and pathological processes, but their detection is difficult, especially in aged and fixed human brain tissue. We used the in-situ proximity ligation assay (PLA) to broaden the range of molecular interactions assessable in-situ in the human neuropathology. We adapted fluorescent in-situ PLA to detect ubiquitin-modified proteins in human brains with Alzheimer’s disease (AD), including approaches for the management of autofluorescence and quantification using a high-content image analysis system. We confirmed that phosphorylated microtubule-associated protein tau (Serine202, Threonine205) aggregates were modified by ubiquitin and that phospho-tau-ubiquitin complexes were increased in hippocampal and frontal cortex regions in AD compared to non-AD brains. Overall, we refined PLA for use in human neuropathology, which has revealed a profound change in the distribution of ubiquitin in AD brain and its association with characteristic tau pathologies.