A synthetic approach for protein evolution and cell engineering

Synthetic biology, such as rewiring cellular signaling circuits by synthetic molecules, allows us to expand our knowledge on dynamic behaviors of biological systems and also develop new biotechnology. Although several researchers have succeeded in engineering cell signaling using unicellular systems such as E.coli or yeast, little has been done using mammalian cells. Here we describe a combinatorial approach for reprogramming cell death (apoptosis) pathways in human cancer cells. To regulate apoptosis pathways in tumor cells, we have established a novel "motif programming" method for generating artificial proteins which modulate cellular signaling. Two motifs respectively related to apoptosis induction and protein transduction were encrypted into different reading frames of an artificial gene (microgene), which was then polymerized; random frame shifts at the junctions between the microgene units yielded combinatorial polymers of three reading frames. We synthesized a bi-functional protein that is able to penetrate through cell membranes and exert a strong apoptotic effect on several cancer cell lines. Because a simple linkage of these motifs was not sufficient to construct a bi-functional peptide, we concluded that the combinatorial strategy is important for reconstituting functions from mixtures of motifs. We also invented a new method for motif programming, and generated cell death modulators from the single motif-mixing library. Rewiring genetic circuits using artificial proteins paves a new way to establish novel pharmaceutical strategies toward cancer treatments