Active expression of the ubiA gene from E. coli in tobacco: Influence of plant ER-Specific signal peptides on the expression of a membrane-bound prenyltransferase in plant cells

The ubiA gene from E. coli codes for 4-hydroxybenzoate: polyprenyldiphospha te 3-polyprenyltransferase, an integral membrane protein involved in ubiqui none biosynthesis. This prokaryotic membrane protein was stably expressed i n tobacco using Agrobacterium tumefaciens-mediated transformation. Transgen ic lines containing a direct fusion of the ubiA structural gene to a 35S-de rived promoter gave very low enzyme activity levels (average 0.16 pkat/mg). Inclusion of an N-terminal ER-specific signal peptide from a lectin gene f rom Phaseolus vulgaris resulted in an average activity of 1.08 pkat/mg in t he transgenic tobacco lines. The additional inclusion of a C-terminal HDEL tetrapeptide, responsible for the retention of proteins in the endoplasmic reticulum of eukaryotic cells, increased the activity to 18.6 pkat/mg. When the promotor of this construct was changed from the 35S derivative to the recently described very strong plant promoter (ocs)(3)mas, the activity inc reased further to 128.6 pkat/mg. The most active tobacco line showed activi ties of the introduced enzyme which exceeded those of wild-type E. coli (th e source of ubiA) by a factor of 1100. These results demonstrate the effica cy of plant ER-specific signal peptides for the active expression of a prok aryotic membrane protein in plants.