In vivo stability of Golgi resident glycolipid glycosyltransferases

SAMPEDRO MARÍA CECILIA; VALDEZ TAUBAS JAVIER; MACCIONI HJF

Mar del Plata, Argentina

Congreso; 43 Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2007

ociedad Argentina de Investigación en Bioquímica y Biología Molecular

The glycolipid glycosyltransferases (GGTs) are N-glycosylated integral membrane proteins of the Golgi complex. Information about the degradation pathways that operate on proteins resident of Golgi complex membranes, in particular the GGTs is lacking. Here we show by pulse chase and Western blot analyses, that the half-life of full length SialT2, GalT2 and GalNAcT stably transfected in CHO-K1 cells, was about 7h. Inhibition of protein synthesis by cycloheximide shortened the half-life to 1,5 h and this decrease was not reverted by inhibitors of lysosomal and proteasomal degradation. Redistribution of these enzymes to the ER by either treating cells with brefeldin A or by impairing the N-glycosylation, lead to their stabilization even in the presence of cycloheximide. Similar results were obtained with constructs in wich the lumenal domain was replaced by GFP, suggesting that degradation of these enzymes is dictated by the N-terminal domain. These results suggest that GGT are degraded by non-classical pathway(s) and that their stability depends on ongoing protein synthesis while in the Golgi but not when re-localised to the ER.