rhodopsins
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Schizorhodopsins (SzRs), a rhodopsin family first identified in Asgard archaea, the archaeal group closest to eukaryotes, are present at a phylogenetically intermediate position between typical microbial rhodopsins and heliorhodopsins. However, the biological function and molecular properties of SzRs have not been reported. Here, SzRs from Asgardarchaeota and from a yet unknown microorganism are expressed in Escherichia coli and mammalian cells, and ion transport assays and patch clamp analyses are used to demonstrate SzR as a novel type of light-driven inward H+ pump. The mutation of a cytoplasmic glutamate inhibited inward H+ transport, suggesting that it functions as a cytoplasmic H+ acceptor. The function, trimeric structure, and H+ transport mechanism of SzR are similar to that of xenorhodopsin (XeR), a light-driven inward H+ pumping microbial rhodopsins, implying that they evolved convergently. The inward H+ pump function of SzR provides new insight into the photobiological life cycle of the Asgardarchaeota.
- MeSH
- Archaea genetika metabolismus MeSH
- buněčná membrána metabolismus MeSH
- fluorescenční protilátková technika MeSH
- gating iontového kanálu účinky záření MeSH
- konformace proteinů MeSH
- molekulární modely MeSH
- multigenová rodina MeSH
- mutace MeSH
- protonové pumpy chemie genetika metabolismus MeSH
- rodopsin chemie genetika metabolismus MeSH
- spektroskopie infračervená s Fourierovou transformací MeSH
- světlo MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The ability to optically image cellular transmembrane voltages at millisecond-timescale resolutions can offer unprecedented insight into the function of living brains in behaving animals. Here, we present a point mutation that increases the sensitivity of Ace2 opsin-based voltage indicators. We use the mutation to develop Voltron2, an improved chemigeneic voltage indicator that has a 65% higher sensitivity to single APs and 3-fold higher sensitivity to subthreshold potentials than Voltron. Voltron2 retained the sub-millisecond kinetics and photostability of its predecessor, although with lower baseline fluorescence. In multiple in vitro and in vivo comparisons with its predecessor across multiple species, we found Voltron2 to be more sensitive to APs and subthreshold fluctuations. Finally, we used Voltron2 to study and evaluate the possible mechanisms of interneuron synchronization in the mouse hippocampus. Overall, we have discovered a generalizable mutation that significantly increases the sensitivity of Ace2 rhodopsin-based sensors, improving their voltage reporting capability.
- MeSH
- akční potenciály fyziologie MeSH
- angiotensin-konvertující enzym 2 * MeSH
- mutace genetika MeSH
- myši MeSH
- neurony fyziologie MeSH
- rodopsin * genetika MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- MeSH
- fotobiologie MeSH
- ještěři MeSH
- peptidy MeSH
- rodopsin fyziologie MeSH
- signální transdukce MeSH
- zevní segment tyčinky metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- srovnávací studie MeSH
Recent advances in phylogenomic analyses and increased genomic sampling of uncultured prokaryotic lineages have brought compelling evidence in support of the emergence of eukaryotes from within the archaeal domain of life (eocyte hypothesis)1,2. The discovery of Asgardarchaeota and its supposed position at the base of the eukaryotic tree of life3,4 provided cues about the long-awaited identity of the eocytic lineage from which the nucleated cells (Eukaryota) emerged. While it is apparent that Asgardarchaeota encode a plethora of eukaryotic-specific proteins (the highest number identified yet in prokaryotes)5, the lack of genomic information and metabolic characterization has precluded inferences about their lifestyles and the metabolic landscape that favoured the emergence of the protoeukaryote ancestor. Here, we use advanced phylogenetic analyses for inferring the deep ancestry of eukaryotes, and genome-scale metabolic reconstructions for shedding light on the metabolic milieu of Asgardarchaeota. In doing so, we: (1) show that Heimdallarchaeia (the closest eocytic lineage to eukaryotes to date) are likely to have a microoxic niche, based on their genomic potential, with aerobic metabolic pathways that are unique among Archaea (that is, the kynurenine pathway); (2) provide evidence of mixotrophy within Asgardarchaeota; and (3) describe a previously unknown family of rhodopsins encoded within the recovered genomes.
- MeSH
- aerobióza MeSH
- anaerobióza MeSH
- Archaea klasifikace genetika metabolismus MeSH
- ekosystém MeSH
- fylogeneze * MeSH
- genom archeí genetika MeSH
- metabolické sítě a dráhy MeSH
- molekulární evoluce MeSH
- rhodopsiny mikrobiální klasifikace genetika MeSH
- RNA ribozomální genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Bacterial proton pumps, proteorhodopsins (PRs), are a major group of light-driven membrane proteins found in marine bacteria. They are functionally and structurally distinct from archaeal and eukaryotic proton pumps. To elucidate the proton transfer mechanism by PRs and understand the differences to nonbacterial pumps on a molecular level, high-resolution structures of PRs' functional states are needed. In this work, we have determined atomic-resolution structures of MAR, a PR from marine actinobacteria, in various functional states, notably the challenging late O intermediate state. These data and information from recent atomic-resolution structures on an archaeal outward proton pump bacteriorhodopsin and bacterial inward proton pump xenorhodopsin allow for deducing key universal elements for light-driven proton pumping. First, long hydrogen-bonded chains characterize proton pathways. Second, short hydrogen bonds allow proton storage and inhibit their backflow. Last, the retinal Schiff base is the active proton donor and acceptor to and from hydrogen-bonded chains.
Microbial heliorhodopsins are a new type of rhodopsins, currently believed to engage in light sensing, with an opposite membrane topology compared to type-1 and type-2 rhodopsins. We determined heliorhodopsins presence/absence is monoderms and diderms representatives from the Tara Oceans and freshwater metagenomes as well as metagenome assembled genome collections. Heliorhodopsins are absent in diderms, confirming our previous observations in cultured Proteobacteria. We do not rule out the possibility that heliorhodopsins serve as light sensors. However, this does not easily explain their absence from diderms. Based on these observations, we speculate on the putative role of heliorhodopsins in light-driven transport of amphiphilic molecules.
- MeSH
- biologické modely MeSH
- gramnegativní bakterie klasifikace genetika MeSH
- metagenom MeSH
- mořská voda mikrobiologie virologie MeSH
- oceány a moře MeSH
- otevřené čtecí rámce MeSH
- senzorické rhodopsiny genetika metabolismus MeSH
- sladká voda mikrobiologie virologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- oceány a moře MeSH
