Commutability of Bilin Chromophores in Plant Phytochromes

Abstract Organisms process environmental light conditions by sensory photoreceptors. In plants, phytochrome photoreceptors detect red and far-red light via covalently bound phytochromobilin (PФB) chromophores to regulate vital adaptive responses including shade avoidance and photomorphogenesis. By contrast, the chromophore of bacterial phytochromes (BphP) is biliverdin (BV), a ubiquitous bilin precursor to PФB biosynthesis. Covalent BV binding by plant phytochromes has not been reported, irrespective of the high structural conservation of the chromophore-binding pockets across the phytochrome superfamily. Unexpectedly, we now find plant phytochromes capable of autocatalytic BV incorporation via the same cysteine residue conventionally used for PФB attachment. BV-bound plant phytochromes retain full functionality as they undergo reversible photoconversion between their Pr and Pfr states and enter light-dependent interactions with downstream partners. BV incorporation into plant phytochromes is inherently inefficient but improves under far-red light and upon introduction of a histidine acting as an acid-base catalyst. Despite these advances, the heterologous deployment of BV-binding plant phytochromes in mammalian cells for optogenetics remains challenging. The chromophore promiscuity evidenced in plant phytochromes extends to BphPs and enables replacing BV by phycocyanobilin (PCB). Bound to PCB, a BphP-based optogenetic circuit retained full functionality but exhibited sensitivity to shorter wavelengths which may benefit application. Our work uncovers an unappreciated commutability of bilin chromophores in the phytochrome superfamily. The position of the active-site cysteine to which the bilin attaches emerges as the major, if not sole, driver for chromophore specificity.