Photomorphogenesis of Myxococcus macrosporus: new insights for light-regulation of cell development.

Myxobacteria are non-photosynthetic bacteria distinguished among prokaryotes by a multicellular stage in their life cycle known as fruiting bodies that are formed in response to nutrient deprivation and stimulated by light. Here, we report an entrained, rhythmic pattern of Myxococcus macrosporus fruiting bodies, forming consistently spaced concentric rings when grown in the dark. Light exposure disrupts this rhythmic phenotype, resulting in a sporadic arrangement and reduced fruiting-body count. M. macrosporus genome encodes a red-light photoreceptor, a bacteriophytochrome (BphP), previously shown to affect the fruiting-body formation in the related myxobacterium Stigmatella aurantiaca. Similarly, the formation of M. macrosporus fruiting bodies is also impacted by the exposure to BphP-specific wavelengths of light. RNA-Seq analysis of M. macrosporus revealed constitutive expression of the bphP gene. Phytochromes, as light-regulated enzymes, control many aspects of plant development including photomorphogenesis. They are intrinsically correlated to circadian clock proteins, impacting the overall light-mediated entrainment of the circadian clock. However, this functional relationship remains unexplored in non-photosynthetic prokaryotes. Genomic analysis unveiled the presence of multiple homologs of cyanobacterial core oscillatory gene, kaiC, in various myxobacteria, including M. macrosporus, S. aurantiaca and M. xanthus. RNA-Seq analysis verified the expression of all kaiC homologs in M. macrosporus and the closely related M. xanthus, which lacks bphP genes. Overall, this study unravels the rhythmic growth pattern during M. macrosporus development, governed by environmental factors such as light and nutrients. In addition, myxobacteria may have a time-measuring mechanism resembling the cyanobacterial circadian clock that links the photoreceptor (BphP) function to the observed rhythmic behavior.

High-resolution crystal structures of a myxobacterial phytochrome at cryo and room temperatures.

Phytochromes (PHYs) are photoreceptor proteins first discovered in plants, where they control a variety of photomorphogenesis events. PHYs as photochromic proteins can reversibly switch between two distinct states: a red light (Pr) and a far-red light (Pfr) absorbing form. The discovery of Bacteriophytochromes (BphPs) in nonphotosynthetic bacteria has opened new frontiers in our understanding of the mechanisms by which these natural photoswitches can control single cell development, although the role of BphPs in vivo remains largely unknown. BphPs are dimeric proteins that consist of a photosensory core module (PCM) and an enzymatic domain, often a histidine kinase. The PCM is composed of three domains (PAS, GAF, and PHY). It holds a covalently bound open-chain tetrapyrrole (biliverdin, BV) chromophore. Upon absorption of light, the double bond between BV rings C and D isomerizes and reversibly switches the protein between Pr and Pfr states. We report crystal structures of the wild-type and mutant (His275Thr) forms of the canonical BphP from the nonphotosynthetic myxobacterium Stigmatella aurantiaca (SaBphP2) in the Pr state. Structures were determined at 1.65 Å and 2.2 Å (respectively), the highest resolution of any PCM construct to date. We also report the room temperature wild-type structure of the same protein determined at 2.1 Å at the SPring-8 Angstrom Compact free electron LAser (SACLA), Japan. Our results not only highlight and confirm important amino acids near the chromophore that play a role in Pr-Pfr photoconversion but also describe the signal transduction into the PHY domain which moves across tens of angstroms after the light stimulus.

Structural basis for light control of cell development revealed by crystal structures of a myxobacterial phytochrome.

Phytochromes are red-light photoreceptors that were first characterized in plants, with homologs in photosynthetic and non-photosynthetic bacteria known as bacteriophytochromes (BphPs). Upon absorption of light, BphPs interconvert between two states denoted Pr and Pfr with distinct absorption spectra in the red and far-red. They have recently been engineered as enzymatic photoswitches for fluorescent-marker applications in non-invasive tissue imaging of mammals. This article presents cryo- and room-temperature crystal structures of the unusual phytochrome from the non-photosynthetic myxo-bacterium Stigmatella aurantiaca (SaBphP1) and reveals its role in the fruiting-body formation of this photomorphogenic bacterium. SaBphP1 lacks a conserved histidine (His) in the chromophore-binding domain that stabilizes the Pr state in the classical BphPs. Instead it contains a threonine (Thr), a feature that is restricted to several myxobacterial phytochromes and is not evolutionarily understood. SaBphP1 structures of the chromophore binding domain (CBD) and the complete photosensory core module (PCM) in wild-type and Thr-to-His mutant forms reveal details of the molecular mechanism of the Pr/Pfr transition associated with the physiological response of this myxobacterium to red light. Specifically, key structural differences in the CBD and PCM between the wild-type and the Thr-to-His mutant involve essential chromophore contacts with proximal amino acids, and point to how the photosignal is transduced through the rest of the protein, impacting the essential enzymatic activity in the photomorphogenic response of this myxobacterium.

Ubiquitous Structural Signaling in Bacterial Phytochromes.

The phytochrome family of light-switchable proteins has long been studied by biochemical, spectroscopic and crystallographic means, while a direct probe for global conformational signal propagation has been lacking. Using solution X-ray scattering, we find that the photosensory cores of several bacterial phytochromes undergo similar large-scale structural changes upon red-light excitation. The data establish that phytochromes with ordinary and inverted photocycles share a structural signaling mechanism and that a particular conserved histidine, previously proposed to be involved in signal propagation, in fact tunes photoresponse.