Evaluation of lettuce chloroplast and soybean cotyledon as platforms for production of functional bone morphogenetic protein 2.

ABSTRACT

The bone morphogenetic protein BMP2 plays a crucial role in the formation and regeneration of bone and cartilage, which is critical for maintaining skeletal integrity and bone fracture repair. Because of its important role in osteogenic properties it has been commercially produced for clinical use. Here we report attempts to express human BMP2 using two plant systems (lettuce chloroplast and soybean seeds). The rhBMP2 gene (coding for the 13 kDa active polypeptide) was introduced in two regions of the lettuce chloroplast genome. Two homoplasmic events were achieved and RT-PCR demonstrated that the BMP2 gene was transcribed. However, it was not possible to detect accumulation of rhBMP2 in leaves. Two soybean events were achieved to express a full-length hBMP2 gene (coding for the 45 kDa pro-BMP2) fused with the α-coixin signal peptide, under control of the ß-conglycinin promoter. Pro-BMP2 was expressed in the transgenic seeds at levels of up to 9.28% of the total soluble seed protein as determined by ELISA. It was demonstrated that this recombinant form was biologically active upon administration to C2C12 cell cultures, because it was able to induce an osteogenic cascade, as observed by the enhanced expression of SP7 (osterix) and ALPI (alkaline phosphatase) genes. Collectively, these results corroborated our previous observation that soybean seeds provide an effective strategy for achieving stable accumulation of functional therapeutic proteins, such as BMP2.