Biosynthesis of N acetyllactosamine glycans in human cell nucleus

PARODI, PEDRO; IRAZOQUI, FERNANDO JOSÉ

Mendoza

Congreso; Congreso SAIB 2022; 2022

SAIB

Oglycosylation is a protein post translational modification in human cells that is critical for cell physiology and involved in severalpathologies In particular, O GlcNAc glycosylation is found in nuclear proteins with regulatory functions in the cell nucleus and showingaltered expression in different cancer types Moreover there is no evidence about the elongation of this glycan in nuclear proteins Inthe present work we study the catalytic activity of β galactosyltransferase in the cell nucleus necessary for the elongation of O GlcNAcglycan in N acetyllactosamine We detected the presence of β 4 Galactosyltransferase 1 in the nucleus of human cells such as CaCo 2HeLa and A 549 by confocal microscopy including z axis cutting This enzyme catalyze the covalent union between galactose usingUDP galactose as donor and O GlcNAc terminals acceptor via β 1  4 bond to form nuclear O acetyllactosamine It is important tomention that UDP Gal is permeable to the nuclear membrane so it is biologically available in the cell nucleus We studied the nuclearβ galactosyltransferase activity through in vitro assays for purified nucleoplasm of different line cells HeLa and CaCo 2 This activitywas measured on different glycoprotein acceptors such as GlcN BSA and ovalbumin and detected by using biotinylated Erythrinacristagalli lectin ( that is able to bond to terminal lactosamine residues In these studies we found an important βgalactosyltransferase activity in cell nucleoplasm We also find constitutive presence of lactosamine residues in nuclear proteins byimmunofluorescence and western blot using ECL as probe Finally we measured lactosamine residues in nuclear proteins such asLamin B 1 and RNA pol 2 by ELISA sandwich assays All together show that the machinery needed for O GlcNAc elongation is located inthe human cell nucleus We are working in additional evidences of this important biochemical discovery