α1,3Fucosyltransferase VI is expressed in HepG2 cells and codistributed with β1,4galactosyltransferase I in the Golgi apparatus and monensin-induced swollen vesicles

The major alpha 1,3fucosyltransferase activity in plasma, liver, and kidney is related to fucosyltransferase VT which is encoded by the FUT6 gene. Here we demonstrate the presence of alpha 1,3fucosyltransferase VI (alpha 3-FucT VI) in the human HepG2 hepatoma cell line by specific activity assays, detection of transcripts, and the use of specific antibodies. First, FucT activity in HepG2 cell lysates was shown to prefer sialyl-N-acetyllactosamine as acceptor substrate indicating expression of alpha 3-FucT VI, RT-PCR analysis further confirmed the exclusive presence of the alpha 3-FucT VI transcripts among the five human alpha 3-FucTs cloned to date, alpha 3-FucT VI was colocalized with beta 1,4galactosyltransferase I(beta 4-GalT I) to the Golgi apparatus by dual confocal immunostaining, Pulse/chase analysis of metabolically labeled alpha 3-FucT VI showed maturation of alpha 3-FucT VI from the early 43 kDa form to the mature, endoglycosidase H-resistant form of 47 kDa which was detected after 2 h of chase. alpha 3-FucT VI was released to the medium and accounted for 50% of overall cell-associated and released enzyme activity. Release occurred by proteolytical cleavage which produced a soluble form of 43 kDa, Monensin treatment segregated alpha 3-FucT VI from the Golgi apparatus to swollen peripheral vesicles where it was colocalized with beta 4-GalT I while alpha 2,6(N)sialyltransferase remained associated with the Golgi apparatus. Both constitutive secretion of alpha 3-FucT VI and its monensin-induced relocation to vesicles analogous to beta 4-GalT I suggest a similar post-Golgi pathway of both alpha 3-FucT VI and beta 4-GalT I.