All it is about location: nuclear O-GalNAc glycosylation

CEJAS, RB; GARAY, YC; IRAZOQUI FJ

Simposio; GlycoAr 2016; 2016

It is well known that polypeptide GalNAc-transferases (ppGalNAc-Ts) initiates O-GalNAc glycosylation of proteins in Golgi apparatus. In the present work we study the O-GalNAc glycosylation of nuclear proteins, modification not described in nucleus at date. Subcellular ppGalNAc-Ts localization was studied both by confocal microscopy and subcellular fractionation. In order to elucidate nuclear O-GalNAc glycosylation by endogenous ppGalNAc-Ts, we purified cellular nucleus that conserves their integrity and are functionally active. Here, we observed the presence of ppGalNAc-T3 isoform in nucleus of several human cell lines and O-GalNAc glycosyl transferase activity in this organelle. UDP-GalNAc was observed in nuclear fraction. Nuclei incubated with sugar-donor UDP-GalNAc raised high level of O-GalNAc protein glycosylation showing an active machinery of nuclear O-GalNAc glycosylation. Nuclear protein glycosylation was analyzed by confocal microscopy, Western Blotting and identified by mass spectrometry. Identified proteins included the Histone H2A, so we performed in vitro histones glycosylation assays and analysis of cellular histone glycosylation by confocal microscopy. Our results demonstrate nuclear O-GalNAc glycosylation of proteins. In a next future, we aim to reveal the cellular significance of this post-translational modification on nuclear proteins.