Growth and differentiation of multicellular systems is orchestrated by spatially restricted gene expression programs in specialized subpopulations. The targeted manipulation of such processes by synthetic tools with high-spatiotemporal resolution could, therefore, enable a deepened understanding of developmental processes and open new opportunities in tissue engineering. Here, we describe the first red/far-red light-triggered gene switch for mammalian cells for achieving gene expression control in time and space. We show that the system can reversibly be toggled between stable on- and off-states using short light pulses at 660 or 740 nm. Red light-induced gene expression was shown to correlate with the applied photon number and was compatible with different mammalian cell lines, including human primary cells. The light-induced expression kinetics were quantitatively analyzed by a mathematical model. We apply the system for the spatially controlled engineering of angiogenesis in chicken embryos. The system’s performance combined with cell- and tissue-compatible regulating red light will enable unprecedented spatiotemporally controlled molecular interventions in mammalian cells, tissues and organisms.

The emergence and future of mammalian synthetic biology depends on technologies for orchestrating and custom tailoring complementary gene expression and signaling processes in a predictable manner. Here, we demonstrate for the first time multi-chromatic expression control in mammalian cells by differentially inducing up to three genes in a single cell culture in response to light of different wavelengths. To this end, we developed an ultraviolet B (UVB)-inducible expression system by designing a UVB-responsive split transcription factor based on the Arabidopsis thaliana UVB receptor UVR8 and the WD40 domain of COP1. The system allowed high (up to 800-fold) UVB-induced gene expression in human, monkey, hamster and mouse cells. Based on a quantitative model, we determined critical system parameters. By combining this UVB-responsive system with blue and red light-inducible gene control technology, we demonstrate multi-chromatic multi-gene control by differentially expressing three genes in a single cell culture in mammalian cells, and we apply this system for the multi-chromatic control of angiogenic signaling processes. This portfolio of optogenetic tools enables the design and implementation of synthetic biological networks showing unmatched spatiotemporal precision for future research and biomedical applications.

We present a synthetic platform based on photo-induced thiol-ene chemistry, by which surfaceattached networks from antimicrobial poly(oxonorbornene) (so-called polymeric synthetic mimics of antimicrobial peptides, SMAMPs) could be easily obtained. By systematically varying hydrophobicity and charge density, surface-attached polymer networks with high antimicrobial activity and excellent cell compatibility were obtained. For the homopolymer networks with constant charge density, antimicrobial activity increased systematically with increasing hydrophobicity (i.e. decreasing swellability and apparent surface energy). Irrespective of charge density, the antimicrobial activity of all networks correlated with the acid constant pK and the isoelectric point (IEP) -the lower pK and IEP, the higher the antimicrobial activity. The cell compatibility of the networks increased with increasing swellability and apparent surface energy, and decreased with increasing charge density. The data corroborates that the mechanism of action of antimicrobial polymer surfaces depends on at least two mechanistic steps, one of which is hydrophobicity-driven and the other charge related. Therefore, we suggest a modified mechanistic model with a charge-driven and a hydrophobicity-driven step. For antimicrobial networks that only varied in hydrophobicity, the antimicrobial activities on surfaces and in solution also correlated -the higher the activity in solution, the higher the activity on surfaces. Thus, the hydrophobicity-driven step for activity on surfaces may be similar to the one in solution. Cell compatibility of SMAMPs in solution and on surfaces also showed a systematic positive correlation for all polymers, therefore this property also depends on the net hydrophobic balance of the polymer.