Near-infrared light is favorable for imaging in mammalian tissues due to low absorbance of hemoglobin, melanin and water. Therefore, fluorescent proteins, biosensors and optogenetic constructs for optimal imaging, optical readout and light manipulation in mammals should have fluorescence and action spectra within the near-infrared window. Interestingly, natural bacterial phytochromes utilize the low molecular weight biliverdin, found in most mammalian tissues, as a photoreactive chromophore. Due to their near-infrared absorbance bacterial phytochromes are preferred templates for designing optical molecular tools for applications in mammals. Based on the analysis of the photochemistry and structure of bacterial phytochromes we suggest a variety of possible bacterial phytochrome-based fluorescent proteins, biosensors, and optogenetic tools. The design strategies and experimental considerations for such probes are proposed. Near-infrared fluorescent proteins and biosensors will extend the methods developed for conventional microscopy into a deep-tissue in vivo macroscopy including multicolor cell and tissue labeling, fluorescence resonance energy transfer, cell photoactivation and tracking, and detection of enzymatic activities and metabolites in tissues. The near-infrared optogenetic tools will allow noninvasive light-control of biochemistry and physiology of a living animal directly through the skin. Moreover, all bacterial phytochromes-based reagents will spectrally complement existing genetically encoded probes in the visible range. The planned probes also will expand our basic knowledge of photochemistry and light-induced signaling of phytochromes in nature. Availability of the near-infrared probes will further stimulate the development of novel in vivo imaging and light-manipulation technologies, optimization of strategies for gene delivery to specific cells and tissues in vivo, design of targeted noninvasive illumination, and refining optical readouts. Overall, this will result in a wide range of noninvasive studies of chemical and metabolic status, as well as molecular and cellular interactions in intact tissues and whole living mammals.
We will develop novel types of fluorescent proteins, biosensors and optogenetic tools for noninvasive imaging, optical detection and light-manipulation of molecular processes and interactions in mammalian cells, tissues and whole model animals. These probes will emit or be activated in near-infrared spectral range, where the tissue transparency is the highest, and will utilize heme-derived biliverdin compound, which is abundant in mammalian tissues, as a chromophore. The proposed near-infrared genetically encoded optical probes will enable visualization, sensing and light-control of biochemistry and physiology of organs and whole organisms in norm and pathology noninvasively through the skin. These proposed probes will also spectrally complement the optical reagents available in the visible range.
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