BETA-1,6 N-ACETYLGLUCOSAMINYLTRANSFERASE (CORE-2 GLCNAC-T) EXPRESSIONIN NORMAL RAT-TISSUES AND DIFFERENT CELL-LINES - EVIDENCE FOR COMPLEXMECHANISMS OF REGULATION

The distribution of the Golgi enzyme pl,beta 1,6-N-acetylglucosaminylt ransferase (core 2 GlcNAc-T for short) has been investigated in severa l tissue and cell systems by combining the potentials of a polyclonal antibody and a novel, sensitive fluorescent enzyme assay. In normal ra t tissues, levels of the protein were found to vary and as a general t rend did not correlate with enzyme activities. Additionally, we observ ed tissue-specific core 2 GlcNAc-T forms of various size: 75 kDa (live r), 70 kDa (spleen), 60 kDA (heart), and 50 kDa (heart and lung). Thes e forms might arise from differential protein modifications; alternati vely, the smaller form may be a product of proteolytic cleavage, given the presence of a catalytically inactive 50 kDa species in rat serum, Chinese hamster ovary (CHO), MDAY-D2, PSA-5E, and PYS-2 cell lines co nsistently displayed a 70 kDa enzyme. When induced to retro-differenti ate in the presence of butyrate + cholera toxin, CHO cells exhibited a 21-fold increase in enzyme activity, while protein levels remained co nstant. A similar trend was observed in the embryonal endoderm cell li nes PSA-5E and PYS-2 where an approximately 100-fold difference in cor e 2 GlcNAc-T activity was found notwithstanding unchanged amounts of t he protein and identical mRNA levels, as evidenced by RT-PCR, In contr ast, levels of core 2 GlcNAc-T activity in MDAY-D2 cells correlated we ll with protein expression. Taken together, these observations demonst rate that core 2 GlcNAc-T expression may be subjected to multiple mech anisms of regulation and suggest that in at least some instances (i,e, , PSA-5E and PYS-2 cells) expression may be regulated exclusively via posttranslational mechanism(s) of control.