Voltage-gated proton channel in a dinoflagellate Smith, Susan M. E; Morgan, Deri; Musset, Boris ...
Proceedings of the National Academy of Sciences - PNAS,
11/2011, Letnik:
108, Številka:
44
Journal Article
Recenzirano
Odprti dostop
Fogel and Hastings first hypothesized the existence of voltage-gated proton channels in 1972 in bioluminescent dinoflagellates, where they were thought to trigger the flash by activating luciferase. ...Proton channel genes were subsequently identified in human, mouse, and Ciona intestinalis, but their existence in dinoflagellates remained unconfirmed. We identified a candidate proton channel gene from a Karlodinium veneficum cDNA library based on homology with known proton channel genes. K. veneficum is a predatory, nonbioluminescent dinoflagellate that produces toxins responsible for fish kills worldwide. Patch clamp studies on the heterologously expressed gene confirm that it codes for a genuine voltage-gated proton channel, kHV1: it is proton-specific and activated by depolarization, its gH–V relationship shifts with changes in external or internal pH, and mutation of the selectivity filter (which we identify as Asp51) results in loss of proton-specific conduction. Indirect evidence suggests that kHV1 is monomeric, unlike other proton channels. Furthermore, kHV1 differs from all known proton channels in activating well negative to the Nernst potential for protons, EH. This unique voltage dependence makes the dinoflagellate proton channel ideally suited to mediate the proton influx postulated to trigger bioluminescence. In contrast to vertebrate proton channels, whose main function is acid extrusion, we propose that proton channels in dinoflagellates have fundamentally different functions of signaling and excitability.
BIOLUMINESCENCE Wilson, Thérèse; Hastings, J. Woodland
Annual review of cell and developmental biology,
01/1998, Letnik:
14, Številka:
1
Journal Article
Recenzirano
Bioluminescence has evolved independently many times; thus the responsible
genes are unrelated in bacteria, unicellular algae, coelenterates, beetles,
fishes, and others. Chemically, all involve ...exergonic reactions of molecular
oxygen with different substrates (luciferins) and enzymes (luciferases),
resulting in photons of visible light ( 50 kcal). In addition to the
structure of luciferan, several factors determine the color of the emissions,
such as the amino acid sequence of the luciferase (as in beetles, for example)
or the presence of accessory proteins, notably GFP, discovered in coelenterates
and now used as a reporter of gene expression and a cellular marker. The
mechanisms used to control the intensity and kinetics of luminescence, often
emitted as flashes, also vary. Bioluminescence is credited with the discovery
of how some bacteria, luminous or not, sense their density and regulate
specific genes by chemical communication, as in the fascinating example of
symbiosis between luminous bacteria and squid.
Bioluminescence Wilson, Thérèse; Hastings, J. Woodland
2013, 2013-02-14
eBook
Bioluminescence is everywhere on earth--most of all in the ocean, from angler fish in the depths to flashing dinoflagellates at the surface. Wilson and Hastings explore the natural history, ...evolution, and biochemistry of the diverse array of organisms that emit light and offer an evolutionary explanation for their sporadic distribution and rarity.
Noctiluca scintillans, a heterotrophic unarmored unicellular bioluminescent dinoflagellate, occurs widely in the oceans, often as a bloom. Molecular phylogenetic analysis based on 18S ribosomal DNA ...sequences consistently has placed this species on the basal branch of dinoflagellates. Here, we report that the structural organization of its luciferase gene is strikingly different from that of the seven luminous species previously characterized, all of which are photosynthetic. The Noctiluca gene codes for a polypeptide that consists of two distinct but contiguous domains. One, which is located in the N-terminal portion, is shorter than but similar in sequence to the individual domains of the three-domain luciferases found in all other luminous dinoflagellates studied. The other, situated in the C-terminal part, has sequence similarity to the luciferin-binding protein of the luminous dinoflagellate Lingulodinium polyedrum, encoded there by a separate gene. Western analysis shows that the native protein has the same size (almost equal to100 kDa) as the heterologously expressed polypeptide, indicating that it is not a polyprotein. Thus, sequences found in two proteins in the L. polyedrum bioluminescence system are present in a single polypeptide in NOCTILUCA:
Enzymes with multiple catalytic sites are rare, and their evolutionary significance remains to be established. This study of luciferases from seven dinoflagellate species examines the previously ...undescribed evolution of such proteins. All these enzymes have the same unique structure: three homologous domains, each with catalytic activity, preceded by an N-terminal region of unknown function. Both pairwise comparison and phylogenetic inference indicate that the similarity of the corresponding individual domains between species is greater than that between the three different domains of each polypeptide. Trees constructed from each of the three individual domains are congruent with the tree of the full-length coding sequence. Luciferase and ribosomal DNA trees both indicate that the Lingulodinium polyedrum luciferase diverged early from the other six. In all species, the amino acid sequence in the central regions of the three domains is strongly conserved, suggesting it as the catalytic site. Synonymous substitution rates also are greatly reduced in the central regions of two species but not in the other five. This lineage-specific difference in synonymous substitution rates in the central region of the domains correlates inversely with the content of GC3, which can be accounted for by the biased usage toward C-ending codons at the degenerate sites. RNA modeling of the central region of the L. polyedrum luciferase domain suggests a function of the constrained synonymous substitutions in the circadian-controlled protein synthesis.
The luciferase of Lingulodinium polyedrum, a marine bioluminescent dinoflagellate, consists of three similar but not identical domains in a single polypeptide. Each encodes an active luciferase that ...catalyzes the oxidation of a chlorophyll-derived open tetrapyrrole (dinoflagellate luciferin) to produce blue light. These domains share no sequence similarity with any other in the GenBank database and no structural or motif similarity with any other luciferase. We report here the 1.8-Å crystal structure of the third domain, D3, at pH 8, and a mechanism for its activity regulation by pH. D3 consists of two major structural elements: a β-barrel pocket putatively for substrate binding and catalysis and a regulatory three-helix bundle. N-terminal histidine residues previously shown to regulate activity by pH are at the interface of the helices in the bundle. Molecular dynamics calculations indicate that, in response to changes in pH, these histidines could trigger a large molecular motion of the bundle, thereby exposing the active site to the substrate.
Hastings comments on an article in a recent issue of PNAS, in which Chun et al. (2004) reported that human respiratory epithelia have the capacity to inactivate a P. aeruginosa quorum-sensing signal. ...This capacity appears to be enzymatic in nature, and it functions in some but not all mammalian cells. This finding opens a new area of research and indicates that humans have evolved mechanisms to interfere with a quorum-sensing pathway.
There is a prominent circadian rhythm of bioluminescence in many species of light-emitting dinoflagellates. In
a daily synthesis and destruction of proteins is used to regulate activity. Experiments ...indicate that the amino acids from the degradation are conserved and incorporated into the resynthesized protein in the subsequent cycle. A different species,
, also exhibits a rhythm of bioluminescence, but the luciferase is not destroyed and resynthesized each cycle. This paper posits that synthesis and destruction constitutes a cellular mechanism to conserve nitrogen in an environment where the resource is limiting.
In this study, the effects of 1 mM sodium nitrite, a reactive nitrogen species (RNS) generator, and 0.5 mM paraquat, which produces reactive oxygen species (ROS), on gene expression in the marine ...dinoflagellate species
Pyrocystis lunula were investigated using microarrays containing 3500 complementary DNAs (cDNAs). A total of 246 differentially expressed genes were identified under these treatments: 204 genes were specifically regulated in response to nitrite and 37 genes specifically to paraquat. Only six genes showed a dependence on both nitrite and paraquat, indicating that the two agents act predominantly via distinct pathways. Although many of these redox-regulated genes encode proteins from a diverse range of functional categories, the majority of them (68%) represent novel sequences. Temporary abnormal spherical cells occurred in nitrite-treated cultures, but not in those exposed to paraquat, suggesting that this response involves a specific pathway triggered by RNS. The genes involved include one that encodes a transcription factor unique to dinoflagellates (HPl), and genes encoding proteins similar to those regulating developmental processes in plants and animals such as NYD-SP5, shaggy and calcium-dependent kinases, the COP9 signalosome complex, ubiquitin-related proteases and a metacaspase.
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