Naslov
Split luciferase NanoLuc as a reporter for the detection of human tau protein oligomerization

Autori
Zubčić, Klara ; Šimić, Goran ; Boban, Mirta

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
9th Croatian Congress of Pharmacology with International Participation / - , 2019, 133-133

Skup
9. hrvatski kongres farmakologije = 9th Croatian Congress of Pharmacology

Mjesto i datum
Zagreb, Hrvatska, 25.09.2019. - 28.09.2019

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
Alzheimer's disease ; luminescent reporter ; neurofibrillary tangles ; protein-protein interactions ; tauopathies ; tau protein oligomerization ; yeast Saccharomyces cerevisiae

Sažetak
One of the main neuropathological hallmarks of Alzheimer's disease are neurofibrillary tangles (NFTs), large aggregates of microtubule binding protein tau that form within the affected neurons. The formation of NFTs is preceded by early-stage tau oligomers, however the molecular pathways that initiate tau aggregation are unclear. To better understand early steps in tau pathology, we constructed a tool for studying tau oligomerization in living cells, based on the luminiscent reporter NanoBiT, in which protein-protein interaction results in the complementation of the luciferase NanoLuc, and consequently in generation of luminescence. Since molecular pathways of protein aggregation are largely evolutionary conserved, we selected a simple cell model, yeast Saccharomyces cerevisiae. We separately fused two luciferase subunits to the human tau protein C-terminus. Expression of tau constructs was verified using western blot. Cells expressing tau separately fused to two luciferase subunits exhibited an increased luminescence, as compared to controls, indicating the formation of tau oligomers. In order to test whether tau-NanoBiT reporter is able to detect sarkosyl-insoluble tau aggregates, we measured tau- NanoBiT luminescence in pho85delta, rim1delta, and sod2delta mutants that were previously reported to have increased levels of sarkosyl-insoluble tau. Our preliminary data showed that the tau-NanoBiT luminescence was elevated in the sod2delta mutant, while the signal in pho85delta and rim1delta mutants was similar to the wild-type levels. In conclusion, we constructed a luminescent reporter for human tau protein oligomerization in living yeast cells. Our future experiments will address whether tau-NanoBiT reporter correlates with the levels of sarkosyl- insoluble tau aggregates.

Izvorni jezik
Engleski

Znanstvena područja
Biologija, Temeljne medicinske znanosti, Kliničke medicinske znanosti, Kognitivna znanost (prirodne, tehničke, biomedicina i zdravstvo, društvene i humanističke znanosti), Biotehnologija u biomedicini (prirodno područje, biomedicina i zdravstvo, biotehničko područje)

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