alpha 1,3Fucosyltransferase VI is expressed in HepG2 cells and codistributed with beta 1,4galactosyltransferase 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. Her e we demonstrate the presence of alpha 1,3fucosyltransferase VI (alpha 3-Fu cT VI) in the human HepG2 hepatoma cell line by specific activity assays, d etection of transcripts, and the use of specific antibodies. First, FucT ac tivity in HepG2 cell lysates was shown to prefer sialyl-N-acetyllactosamine as acceptor substrate indicating expression of alpha 3-FucT VI, RT-PCR ana lysis further confirmed the exclusive presence of the alpha 3-FucT VI trans cripts among the five human alpha 3-FucTs cloned to date, alpha 3-FucT VI w as colocalized with beta 1,4galactosyltransferase I(beta 4-GalT I) to the G olgi apparatus by dual confocal immunostaining, Pulse/chase analysis of met abolically labeled alpha 3-FucT VI showed maturation of alpha 3-FucT VI fro m 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 fr om the Golgi apparatus to swollen peripheral vesicles where it was colocali zed with beta 4-GalT I while alpha 2,6(N)sialyltransferase remained associa ted with the Golgi apparatus. Both constitutive secretion of alpha 3-FucT V I 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-G alT I.