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Evolutionary steps involving counterion displacement in a tunicate opsin.

ABSTRACT: Ci-opsin1 is a visible light-sensitive opsin present in the larval ocellus of an ascidian, Ciona intestinalis This invertebrate opsin belongs to the vertebrate visual and nonvisual opsin groups in the opsin phylogenetic tree. Ci-opsin1 contains candidate counterions (glutamic acid residues) at positions 113 and 181; the former is a newly acquired position in the vertebrate visual opsin lineage, whereas the latter is an ancestral position widely conserved among invertebrate opsins. Here, we show that Glu113 and Glu181 in Ci-opsin1 act synergistically as counterions, which imparts molecular properties to Ci-opsin1 intermediate between those of vertebrate- and invertebrate-type opsins. Synergy between the counterions in Ci-opsin1 was demonstrated by E113Q and E181Q mutants that exhibit a pH-dependent spectral shift, whereas only the E113Q mutation in vertebrate rhodopsin yields this spectral shift. On absorbing light, Ci-opsin1 forms an equilibrium between two intermediates with protonated and deprotonated Schiff bases, namely the MI-like and MII-like intermediates, respectively. Adding G protein caused the equilibrium to shift toward the MI-like intermediate, indicating that Ci-opsin1 has a protonated Schiff base in its active state, like invertebrate-type opsins. Ci-opsin1's G protein activation efficiency is between the efficiencies of vertebrate- and invertebrate-type opsins. Interestingly, the E113Y and E181S mutations change the molecular properties of Ci-opsin1 into those resembling invertebrate-type or bistable opsins and vertebrate ancient/vertebrate ancient-long or monostable opsins, respectively. These results strongly suggest that acquisition of counterion Glu113 changed the molecular properties of visual opsin in a vertebrate/tunicate common ancestor as a crucial step in the evolution of vertebrate visual opsins.

SUBMITTER: Kojima K

PROVIDER: S-EPMC5468630 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Evolutionary steps involving counterion displacement in a tunicate opsin.

Kojima Keiichi K Yamashita Takahiro T Imamoto Yasushi Y Kusakabe Takehiro G TG Tsuda Motoyuki M Shichida Yoshinori Y

Proceedings of the National Academy of Sciences of the United States of America 20170522 23

Ci-opsin1 is a visible light-sensitive opsin present in the larval ocellus of an ascidian, <i>Ciona intestinalis</i> This invertebrate opsin belongs to the vertebrate visual and nonvisual opsin groups in the opsin phylogenetic tree. Ci-opsin1 contains candidate counterions (glutamic acid residues) at positions 113 and 181; the former is a newly acquired position in the vertebrate visual opsin lineage, whereas the latter is an ancestral position widely conserved among invertebrate opsins. Here, w ...[more]

PMID: 28533401

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Project description:BackgroundAnalyzing close species with diverse developmental modes is instrumental for investigating the evolutionary significance of physiological, anatomical and behavioral features at a molecular level. Many examples of trait loss are known in metazoan populations living in dark environments. Tunicates are the closest living relatives of vertebrates and typically present a lifecycle with distinct motile larval and sessile adult stages. The nervous system of the motile larva contains melanized cells associated with geotactic and light-sensing organs. It has been suggested that these are homologous to vertebrate neural crest-derived melanocytes. Probably due to ecological adaptation to distinct habitats, several species of tunicates in the Molgulidae family have tailless (anural) larvae that fail to develop sensory organ-associated melanocytes. Here we studied the evolution of Tyrosinase family genes, indispensible for melanogenesis, in the anural, unpigmented Molgula occulta and in the tailed, pigmented Molgula oculata by using phylogenetic, developmental and molecular approaches.ResultsWe performed an evolutionary reconstruction of the tunicate Tyrosinase gene family: in particular, we found that M. oculata possesses genes predicted to encode one Tyrosinase (Tyr) and three Tyrosinase-related proteins (Tyrps) while M. occulta has only Tyr and Tyrp.a pseudogenes that are not likely to encode functional proteins. Analysis of Tyr sequences from various M. occulta individuals indicates that different alleles independently acquired frameshifting short indels and/or larger mobile genetic element insertions, resulting in pseudogenization of the Tyr locus. In M. oculata, Tyr is expressed in presumptive pigment cell precursors as in the model tunicate Ciona robusta. Furthermore, a M. oculata Tyr reporter gene construct was active in the pigment cell precursors of C. robusta embryos, hinting at conservation of the regulatory network underlying Tyr expression in tunicates. In contrast, we did not observe any expression of the Tyr pseudogene in M. occulta embryos. Similarly, M. occulta Tyr allele expression was not rescued in pigmented interspecific M. occulta × M. oculata hybrid embryos, suggesting deleterious mutations also to its cis-regulatory sequences. However, in situ hybridization for transcripts from the M. occulta Tyrp.a pseudogene revealed its expression in vestigial pigment cell precursors in this species.ConclusionsWe reveal a complex evolutionary history of the melanogenesis pathway in tunicates, characterized by distinct gene duplication and loss events. Our expression and molecular data support a tight correlation between pseudogenization of Tyrosinase family members and the absence of pigmentation in the immotile larvae of M. occulta. These results suggest that relaxation of purifying selection has resulted in the loss of sensory organ-associated melanocytes and core genes in the melanogenesis biosynthetic pathway in M. occulta.

Project description:Experimental studies of primordial metabolic evolution are based on multi-component reactions which typically result in highly complex product mixtures. The detection and structural assignment of these products crucially depends on sensitive and selective analytical procedures. Progress in the instrumentation of these methods steadily lowered the detection limits to concentrations in the pico molar range. At the same time, conceptual improvements in chromatography, nuclear magnetic resonance (NMR) and mass spectrometry dramatically increased the resolution power as well as throughput, now, allowing the simultaneous detection and structural determination of hundreds to thousands of compounds in complex mixtures. In retrospective, the development of these analytical methods occurred stepwise in a kind of evolutionary process that is reminiscent of steps occurring in the evolution of metabolism under chemoautotrophic conditions. This can be nicely exemplified in the analytical procedures used in our own studies that are based on Wächtershäuser's theory for metabolic evolution under Fe/Ni-catalyzed volcanic aqueous conditions. At the onset of these studies, gas chromatography (GC) and GC-MS (mass spectrometry) was optimized to detect specific low molecular weight products (<200 Da) in a targeted approach, e.g., methyl thioacetate, amino acids, hydroxy acids, and closely related molecules. Liquid chromatography mass spectrometry (LC-MS) was utilized for the detection of larger molecules including peptides exceeding a molecular weight of 200 Da. Although being less sensitive than GC-MS or LC-MS, NMR spectroscopy benefitted the structural determination of relevant products, such as intermediates involved in a putative primordial peptide cycle. In future, Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) seems to develop as a complementary method to analyze the compositional space of the products and reaction clusters in a non-targeted approach at unprecedented sensitivity and mass resolution (700,000 for m/z 250). Stable isotope labeling was important to differentiate between reaction products and artifacts but also to reveal the mechanisms of product formation. In this review; we summarize some of the developmental steps and key improvements in analytical procedures mainly used in own studies of metabolic evolution.

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Evolutionary steps involving counterion displacement in a tunicate opsin.

Publications

Evolutionary steps involving counterion displacement in a tunicate opsin.

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