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Quorum sensing is a cell-to-cell communication system used by pathogenic bacteria to control expression of virulence factors. In Pseudomonas aeruginosa, quorum-sensing mutants show reduced virulence and, it is 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.

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. [ABSTRACT FROM AUTHOR]

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 tetra- pyrrole (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. [ABSTRACT FROM AUTHOR]

Although nucleosomes and histones are lacking in dinoflagellate nuclei, small basic histone-like proteins have been reported, but their function(s) is unknown. In this study we cloned and sequenced a gene for a histone-like protein from the dinoflagellate Lingulodinium polyedrum (Stein) Dodge (HLp) (formerly Gonyaulax polyedra Stein) and investigated its post-translational modification and DNA-binding activities. HLp appears to be acetylated in L. polyedrum , and we identified several L. polyedrum proteins that possess histone acetyltransferase activity and may be responsible for this modification. HLp binds weakly to L. polyedrum DNA but to certain specific sequences with higher affinity, consistent with its having a regulatory function. [ABSTRACT FROM AUTHOR]

Examines the difference of the members of a dinoflagellate luciferase gene family in synonymous substitution rates. Exhibition of unusual structure and organization for Lingulodinium polyedrum genes; Indications of high interspecific conservation between the domains of P. lunula and P. polyedrum; Difference between the genes of P. lunula and P. polyedrum.

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 (approximately 100 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.

Both pulsed and continuous applications of the RNA polymerase II inhibitor thiolutin cause a dramatic but reversible loss of bioluminescence and its overt rhythmicity in cells of the dinoflagellate Lingulodinium polyedrum (formerly Gonyaulax polyedra). Such cells remain alive, and the rhythm resumes after an interval, the length of which depends on the concentration of thiolutin used. The period and phase of the resumed rhythm were not systematically altered following such treatments, and the effects were not different at different circadian phases. For three different genes, luciferin binding protein (lbp), luciferase (lcf), and glyceraldehyde-3-phosphate dehydrogenase (gapdh), which are circadian-regulated at the level of translation, the amounts of their mRNAs were determined by Northern blots for times up to 12.5 h following the addition of 1.5 microM thiolutin. Consistent with previous reports that their abundances do not change with circadian time, their levels remained high for several hours after thiolutin addition, but then did diminish.

Lingulodinium polyedrum luciferase is a bioluminescent protein found in the marine dinoflagellate formerly known as Gonyaulax. It is located in organelles called scintillons that emit brief and bright flashes of light that are regulated by an endogenous circadian clock. The complete luciferase molecule has a molecular mass of 136 994 Da and contains three homologous domains, each of which is a separately active luciferase. Two of these domains, D2-LCF and D3-LCF, have been cloned, expressed and crystallized. Crystals of D2-LCF were obtained from PEG 10 000 in space group P2(1)2(1)2(1), with unit-cell parameters a = 49.1, b = 104.7, c = 180.3 A. They diffract to 2.9 A on a rotating anode. Crystals of D3-LCF were grown from PEG 2000 in space group P2(1)2(1)2(1), with unit-cell parameters a = 58.86, b = 63.98, c = 95.76 A. They diffract to 2.3 A on a rotating anode.