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2004 Funded Pilot Projects
Characterization of a cyanobacterial anti-algal compound
Eric Webb and Chris Reddy, Woods Hole Oceanographic Institution
Cnidarian toxins against voltage-gated Ca2+ channelsRobert Greenberg, Marine Biological Laboratory
Cnidarians such as jellyfish and sea anemones produce venoms that are comprised of a variety of toxins. Several of these toxins have been characterized and are targeted against specific receptors and ion channels in excitable cells. For example, polypeptide sea anemone toxins have dramatic effects on voltage-gated sodium channels. They bind selectively to these channels and inhibit inactivation of the channel.
Peptide toxins from sea anemones have also been shown to selectively inhibit different types of voltage-gated potassium channels. Voltage-gated calcium (Ca2+) channels are critical components of excitable cells. They open in response to changes in membrane potential and regulate levels of intracellular Ca2+, an important second messenger. Ca2+ channels are targets for several agents used to treat cardiovascular diseases, and they are also being exploited as targets for pharmacological management of pain, epilepsy, and neural ischemia and stroke, as well as other conditions.
Ca2+ channels are also acted upon by toxins from arthropods and molluscs. Many cnidarian venoms serve to paralyze their prey, and Ca2+ channels associated with the neuromuscular junction may be specifically targeted by toxins found within these venoms.
This project will use heterologously expressed Ca2+ channels to screen cnidarian venoms for toxins that interact with these channels. Various species of cnidarians will be collected from east-coast waters, and crude venom extracts will be tested for their effects on expressed Ca2+ currents. If specific effects are found, the active toxin components will be isolated by standard biochemical approaches (eg, HPLC), and characterized. These experiments may provide new pharmacological tools for analysis of these channels, and may eventually lead to novel drugs for therapeutic use.
Marine phage as vectors of gene transfer between marine bacteria and bacterial pathogensJon King, Massachusetts Institute of Technology
Bacterial infections continue to be a major source of disease and mortality worldwide. A diverse set of genes and gene clusters necessary for bacterial pathogenesis have been carefully documented in the last decade. A subset of these virulence genes are found encoded within facultative pathogens by bacteriophages in the integrated, or lysogenic state (prophages). Conversion of avirulent bacteria to virulent forms by phage lysogeny has been demonstrated in the laboratory and inferred through genomic comparisons between closely related bacterial strains differing in pathogenicity. However, very little is known of the sources and environmental distribution of phage-encoded virulence genes or the evolutionary routes through which they have been acquired by their bacterial hosts.
Bacteriophages are extremely abundant in marine waters, with numbers approaching 107/ml. Recent genomic data suggest that phages of diverse hosts can exchange sequences within a global gene pool. This exchange is reflected in the surprising number of genetic and structural features common between phages of marine heterotrophic bacteria, phages of marine photosynthetic bacteria, and well studied phages of enteric bacteria, including the types causing human diseases. Therefore, the ocean may be an important reservoir of phageencoded virulence genes and may be an active site for phage mediated exchange of these sequences between bacterial populations.
The overall goals of this project are to investigate whether phage-borne genes or gene clusters associated with virulence effects in bacterial pathogens of humans, are also found among phages infecting marine photosynthetic bacteria.
The experiments involve:
a) Isolation of phages of marine and enteric bacteria from estuaries and coastal waters where enteric bacteria from terrestrial runoff or sewage effluents mix with coastal ocean waters carrying marine bacteria. b) Using oligonucleotide probes for known phage-borne virulence genes, assaying filtered water samples for these sequences. If detected, further investigation would determine whether they were associated with isolatable phages of either laboratory enteric bacteria and/or coastal ecotypes of the marine obligate photoautoroph, Synechococcus.
Positive results would open the possibility of identifying sources of extant as well as emerging pathogenic sequences through more systematic investigation of marine microorganisms.