Characterization of bovine core 2 beta-1,6-N-acetylglucosaminyltransferase-M gene, tissue-specific expression, and herpes virus-4 homologue

Mucins are large molecular weight glycoproteins found in mucus secretion and cell surface. The biological functions of mucins primarily reside in the O-glycans, which constitute 70--80% of mucin molecule by weight and are very heterogeneous. The complexity of mucin O-glycans can be further elaborated by branching enzymes, including core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT)-1(L), -2(M), and -3(T), and blood group I GnT. C2GnT activity has been detected in broad spectrum of tissues yet expression of the three C2GnT genes is specific to cell type, and physiological and pathological conditions. The major focus of this dissertation is characterization of tissue-specific transcripts of bovine C2GnT-M and its genomic structure. Four different bovine C2GnT-M transcripts were identified and their tissue-specific expression profile was examined. Two bC2GnT-M transcripts were found exclusively in tracheal epithelium and testis, whereas the other two were found in other mucus-secreting tissues. The bC2GnT-M gene contains four exons spanning 5.3 kb, and the entire open reading frame is in one exon. A bovine viral C2GnT-M gene was also identified. The viral C2GnT-M is the only known C2GnT gene in virus and it is homologous with bC2GnT-M. The similarity of genomic structures of bovine and viral C2GnT-Ms suggests that the viral C2GnT-M gene is originated from bovine host. Second part of this dissertation will describe development of a mouse embryonic stem cell line that lacks a copy of C2GnT-L gene. The ORF of C2GnT-L gene was replaced with that of a green fluorescence protein gene in this ES cells. Mice to be developed from the ES cells may be used to study the expression of C2GnT-L gene in animals under various physiological and pathological conditions. In a separate project, effect of epigenetic modification on expression of CMV-promoter-driven transgene was examined. Inhibitors for histone deacetylase or DNA methylase was used to show treatment of these drugs can enhance the activity of CMV-promoter.