David Morse

Heavy metal-induced oxidative stress in algae

Heavy metals, depending on their oxidation states, can be highly reactive and, as a consequence, toxic to most organisms. They are produced by an expanding variety of anthropogenic sources suggesting an increasingly important role for this form of pollution. The toxic effect of heavy metals appears to be related to production of reactive oxygen species (ROS) and the resulting unbalanced cellular redox status. Algae respond to heavy metals by induction of several antioxidants, including diverse enzymes such as superoxide dismutase, catalase, glutathione peroxidase and ascorbate peroxidase, and the synthesis of low molecular weight compounds such as carotenoids and glutathione. At high, or acute, levels of metal pollutants, damage to algal cells occurs because ROS levels exceed the capacity of the cell to cope. At lower, or chronic, levels algae accumulate heavy metals and can pass them on to organisms of other trophic levels such as mollusks, crustaceans, and fishes. We review here the evidence linking metal accumulation, cellular toxicity, and the generation of ROS in aquatic environments.

S-RNase uptake by compatible pollen tubes in gametophytic self-incompatibility.

Many flowering plants avoid inbreeding through a genetic mechanism termed self-incompatibility. An extremely polymorphic S-locus controls the gametophytic self-incompatibility system that causes pollen rejection (that is, active arrest of pollen tube growth inside the style) when an S-allele carried by haploid pollen matches one of the S-alleles present in the diploid style. The only known product of the S-locus is an S-RNase expressed in the mature style. The pollen component to this cell–cell recognition system is unknown and current models propose that it either acts as a gatekeeper allowing only its cognate S-RNase to enter the pollen tube, or as an inhibitor of non-cognate S-RNases. In the latter case, all S-RNases are presumed to enter pollen tubes; thus, the two models make diametrically opposed predictions concerning the entry of S-RNases into compatible pollen. Here we use immunocytochemical labelling of pollen tubes growing in styles to show accumulation of an S-RNase in the cytoplasm of all pollen-tube haplotypes, thus providing experimental support for the inhibitor model.

Two circadian oscillators in one cell

A CIRCADIAN clock, which continues to oscillate in constant conditions, is almost ubiquitous in eukaryotes as well as some prokaryotes1. This class of biological oscillators drives daily rhythms as diverse as photosynthesis in plants2 and the sleep-wake cycle in man3 and enables organisms to anticipate environmental changes or segregate in time-incompatible processes4. Circadian oscillators share many properties, suggesting that the clock is a single mechanism, preserved throughout evolution, which is capable of controlling all the different circadian functions. Here we show that two rhythms in a unicellular organism can, under certain experimental conditions, run independently, and thus each rhythm must be controlled by its own distinct oscillator.

Hypervariable Domains of Self-incompatibility RNases Mediate Allele-Specific Pollen Recognition

Self-incompatibility (SI) in angiosperms is a genetic mechanism that promotes outcrossing through rejection of self-pollen. In the Solanaceae, SI is determined by a multiallelic S locus whose only known product is an S RNase. S RNases show a characteristic pattern of five conserved and two hypervariable regions. These are thought to be involved in the catalytic function and in allelic specificity, respectively. When the Solanum chacoense S12S14 genotype is transformed with an S11 RNase, the styles of plants expressing significant levels of the transgene reject S11 pollen. A previously characterized S RNase, S13, differs from the S11 RNase by only 10 amino acids, four of which are located in the hypervariable regions. When S12S14 plants were transformed with a chimeric S11 gene in which these four residues were substituted with those present in the S13 RNase, the transgenic plants acquired the S13 phenotype. This result demonstrates that the S RNase hypervariable regions control allelic specificity.

Phenotypic Rescue of a Peripheral Clock Genetic Defect via SCN Hierarchical Dominance

The mammalian circadian system contains both central and peripheral oscillators. To understand the communication pathways between them, we have studied the rhythmic behavior of mouse embryo fibroblasts (MEFs) surgically implanted in mice of different genotypes. MEFs from Per1(-/-) mice have a much shorter period in culture than do tissues in the intact animal. When implanted back into mice, however, the Per1(-/-) MEF take on the rhythmic characteristics of the host. A functioning clock is required for oscillations in the target tissues, as arrhythmic clock(c/c) MEFs remain arrhythmic in implants. These results demonstrate that SCN hierarchical dominance can compensate for severe intrinsic genetic defects in peripheral clocks, but cannot induce rhythmicity in clock-defective tissues.

The Symbiodinium kawagutii genome illuminates dinoflagellate gene expression and coral symbiosis

Symbionts are adapted to work with corals Many corals have formed mutualistic associations with dinoflagellate symbionts, which are thought to provide nutrients and other benefits. To examine the underlying genetics of this association, S. Lin et al. sequenced the genome of the endosymbiont dinoflagellate Symbiodinium kawagutii. The genome includes gene number expansions and encodes microRNAs that show complementarity to genes within the coral genome. Such microRNAs may be involved in regulating coral genes. Furthermore, coral and S. kawagutii appear to share homologs of genes encoding specific nutrient transporters. The findings shed light on how symbiosis is established and maintained between dinoflagellates and corals. Science, this issue p. 691 The genome of the coral symbiont Symbiodinium reveals fundamental aspects of the coral-alga symbiosis. Dinoflagellates are important components of marine ecosystems and essential coral symbionts, yet little is known about their genomes. We report here on the analysis of a high-quality assembly from the 1180-megabase genome of Symbiodinium kawagutii. We annotated protein-coding genes and identified Symbiodinium-specific gene families. No whole-genome duplication was observed, but instead we found active (retro)transposition and gene family expansion, especially in processes important for successful symbiosis with corals. We also documented genes potentially governing sexual reproduction and cyst formation, novel promoter elements, and a microRNA system potentially regulating gene expression in both symbiont and coral. We found biochemical complementarity between genomes of S. kawagutii and the anthozoan Acropora, indicative of host-symbiont coevolution, providing a resource for studying the molecular basis and evolution of coral symbiosis.

Production of an S RNase with Dual Specificity Suggests a Novel Hypothesis for the Generation of New S Alleles

Gametophytic self-incompatibility in plants involves rejection of pollen when pistil and pollen share the same allele at the S locus. This locus is highly multiallelic, but the mechanism by which new functional S alleles are generated in nature has not been determined and remains one of the most intriguing conceptual barriers to a full understanding of selfincompatibility. The S11 and S13 RNases of Solanum chacoense differ by only 10 amino acids, but they are phenotypically distinct (i.e., they reject either S11 or S13 pollen, respectively). These RNases are thus ideally suited for a dissection of the elements involved in recognition specificity. We have previously found that the modification of four amino acid residues in the S11 RNase to match those in the S13 RNase was sufficient to completely replace the S11 phenotype with the S13 phenotype. We now show that an S11 RNase in which only three amino acid residues were modified to match those in the S13 RNase displays the unprecedented property of dual specificity (i.e., the simultaneous rejection of both S11 and S13 pollen). Thus, S12S14 plants expressing this hybrid S RNase rejected S11, S12, S13, and S14 pollen yet allowed S15 pollen to pass freely. Surprisingly, only a single base pair differs between the dual-specific S allele and a monospecific S13 allele. Dual-specific S RNases represent a previously unsuspected category of S alleles. We propose that dualspecific alleles play a critical role in establishing novel S alleles, because the plants harboring them could maintain their old recognition phenotype while acquiring a new one.

Plastid ultrastructure defines the protein import pathway in dinoflagellates

Eukaryotic cells contain a variety of different compartments that are distinguished by their own particular function and characteristic set of proteins. Protein targeting mechanisms to organelles have an additional layer of complexity in algae, where plastids may be surrounded by three or four membranes instead of two as in higher plants. The mechanism of protein import into dinoflagellates plastids, however, has not been previously described despite the importance of plastid targeting in a group of algae responsible for roughly half the oceans net primary production. Here, we show how nuclear-encoded proteins enter the triple membrane-bound plastids of the dinoflagellate Gonyaulax. These proteins all contain an N-terminal leader sequence with two distinct hydrophobic regions flanking a region rich in hydroxylated amino acids (S/T). We demonstrate that plastid proteins transit through the Golgi in vivo, that the first hydrophobic region in the leader acts as a typical signal peptide in vitro, and that the S/T-rich region acts as a typical plastid transit sequence in transgenic plants. We also show that the second hydrophobic region acts as a stop transfer sequence so that plastid proteins in Golgi-derived vesicles are integral membrane proteins with a predominant cytoplasmic component. The dinoflagellate mechanism is thus different from that used by the phylogenetically related apicomplexans, and instead, is similar to that of the phylogenetically distant Euglena, whose plastids are also bound by three membranes. We conclude that the protein import mechanism is dictated by plastid ultrastructure rather than by the evolutionary history of the cell.

Time after time: inputs to and outputs from the mammalian circadian oscillators

Oscillating levels of clock gene transcripts in the suprachiasmatic nucleus (SCN) are essential components of the mammalian circadian pacemaker. Their synchronization with daily light cycles involves neural connections from light-sensitive photoreceptor-containing retinal ganglion cells. This clock orchestrates rhythmic expression for approximately 10% of the SCN gene transcripts, of which only 10% are also rhythmically expressed in other tissues. Many of the transcripts expressed rhythmically only in the SCN are involved in neurosecretion, and their secreted products could mediate SCN control over physiological rhythms by coordinating rhythmicity in other nuclei within the brain. The coordination of clock gene transcript oscillations in peripheral tissues could be controlled directly by specific signals or indirectly by rhythmic behavior such as feeding.

Structure and organization of the peridinin-chlorophyll a-binding protein gene in Gonyaulax polyedra

Abstract We have identified a major 32-kDa protein in the dinoflagellate Gonyaulax polyedra as a peridinin-chlorophyll a-binding protein (PCP), based on microsequence data and immunological cross-reaction with antibodies raised against PCP from another dinoflagellate species. A cDNA for this protein, identified by a PCR-based cloning strategy, encoded all 68 of the amino acids microsequenced, thus confirming the identity of the clone. The PCP gene is highly expressed at both the mRNA and protein levels, and only PCP transcripts corresponding in size to the cDNA sequence were detected. Slot blot analyses show that there are roughly 5000 copies of the PCP gene in Gonyaulax, making this gene one of the most highly repeated protein-coding genes ever reported, yet the sequence of the different gene copies in the genome appears extraordinarily well conserved as judged by Southern blot analyses. The gene, as indicated by Southern blot and PCR data, is suggested to be present in 5000 intronless copies arranged head to tail in the genome, separated by conserved 1-kb spacers. Based on the conserved sequence of the spacer region, its presence next to each of the PCP coding sequences, and the uniform size of the PCP transcript, we propose that this region represents a dinoflagellate transcriptional promoter. This putative promoter region contains none of the sequence elements for DNA-binding proteins involved in transcriptional initiation reported in other organisms.