Overexpression of human erythropoietin in tobacco does not affect plant fertility or morphology

Human erythropoietin (EPO) is a leading product in the biopharmaceutical market, but functional EPO has only been produced in mammalian cells, which limits its application and drives up the production costs. Using plants to produce human proteins may be an alternative way to reduce the cost. However, a recent report demonstrated that overexpression of the human EPO gene (EPO) in tobacco or Arabidopsis rendered males sterile and retarded vegetative growth, which raises concern whether EPO might interfere with hormone levels in transgenic plants. In the present study, we demonstrated that overexpressing EPO with additional 5'-His tag and 3' ER-retention peptides in tobacco did not cause any developmental defect compared to GUS plants. With our method, all 20 transgenic plants grew on selective medium and, further confirmed by PCR, were fertile. Most of them grew similarly compared to GUS plants. Only one transgenic plant (EPO2) was shorter in plant height but had twice the life span compared to other transgenic plants. When 11 randomly selected EPO plants, along with the abnormal plant EPO2, were subjected to RT-PCR analysis, all of them had detectable EPO transcripts. However, their protein levels varied considerably; seven of them had detectable EPO proteins analyzed by western blot. Our results indicate that overexpressing human EPO protein in plants does not have detrimental effects on growth and development. Our transformation systems allow us to further explore the possibility of glycoengineering tobacco plants for producing functional EPO and its derivatives.