Phenotypically Complex Living Materials Containing Engineered Cyanobacteria

Abstract

A cyanobacterial photosynthetic biocomposite material was fabricated using 3D-printing and bioengineered to produce multiple functional outputs in response to an external chemical stimulus. Our investigations show the advantages of utilizing additive manufacturing techniques in controlling the design and shape of the fabricated materials, which proved to be important for the support and growth of obligate phototrophic microorganisms within the material. As an initial proof-of-concept, a synthetic theophylline-responsive riboswitch in Synechococcus elongatus PCC 7942 was used for regulating the expression of a yellow fluorescent protein (YFP) reporter. Upon induction with theophylline, the encapsulated cells produced YFP within the hydrogel matrix. Subsequently, a strain of S. elongatus was engineered to produce an oxidative enzyme that is useful for bioremediation, laccase, expressed either constitutively or under the control of the riboswitch. The responsive biomaterial can decolorize a common textile dye pollutant, indigo carmine, potentially serving as a useful tool in environmental bioremediation. Finally, cells were engineered to have the capacity for inducible cell death to eliminate their presence once their activity is no longer required, which is an important function for biocontainment and minimizing unintended environmental impact. By integrating genetically engineered stimuli-responsive cyanobacteria in patterned volumetric 3D-printed designs, we demonstrate the potential of programmable photosynthetic biocomposite materials capable of producing functional outputs including, but not limited to, bioremediation.