Developing Sugarcane As a Bio-Factory for High Level Production of a Potent Recombinant Anti-Viral Protein

The increasing demand for high-value proteins requires the development of new expression platforms that results in high yields. Expression of these proteins in plants represents a promising alternative, especially for complex proteins and glycoproteins that are difficult to produce in other systems. The use of plants as bio-factories has gained a lot of attention due to high scalability, low cost and low risk of mammalian contaminants. Sugarcane is an ideal expression platform due to its vegetative propagation, efficient photosynthesis, high biomass production capacity and rapid growth rate. The objective of this project is to develop sugarcane as an efficient expression system for the production of large quantities of a high-value protein with potent anti-viral and anti-tumor properties. Our previous work yielded up to 0.1% total soluble protein (TSP, equivalent to 0.72mg/kg fresh weight) in sugarcane lines expressing the protein under the control of the maize Ubiquitin (Ubi1) constitutive promoter. In an effort to increase yield, the therapeutic protein was co-expressed from multiple promoters developed in our laboratory, such as the stem-regulated promoters from Sugarcane DIRIGENT5-1 (SHDIR5-1 ) and DIRIGENT16 (SHDIR16), and the constitutive promoters Sugarcane bacilliform virus (SCBV21), Sugarcane ELONGATION FACTOR1a (SHEF1a), and PROLINE RICH PROTEIN (SHPRP). Protein yields were significantly increased from 0.1% to 0.8% TSP (5.99 mg/kg stalk fresh weight) on transgenic lines carrying the co-expressed protein under the control of Ubi1, SCBV21 and SHDIR5-1 promoters. Yields of the therapeutic protein recovered with expression from multiple promoters represent an 8-fold increase from those obtained with expression from a single promoter.