title: Subcellular localization of dinoflagellate polyketide synthases and fatty acid synthase activity.
abstract: Dinoflagellates are prolific producers of polyketide secondary metabolites. Dinoflagellate polyketide synthases (PKSs) have sequence similarity to Type I PKSs, megasynthases that encode all catalytic domains on a single polypeptide. However, in dinoflagellate PKSs identified to date, each catalytic domain resides on a separate transcript, suggesting multiprotein complexes similar to Type II PKSs. Here, we provide evidence through coimmunoprecipitation that single-domain ketosynthase and ketoreductase proteins interact, suggesting a predicted multiprotein complex. In Karenia brevis (C.C. Davis) Gert Hansen & Ø. Moestrup, previously observed chloroplast localization of PKSs suggested that brevetoxin biosynthesis may take place in the chloroplast. Here, we report that PKSs are present in both cytosol and chloroplast. Furthermore, brevetoxin is not present in isolated chloroplasts, raising the question of what chloroplast-localized PKS enzymes might be doing. Antibodies to K. brevis PKSs recognize cytosolic and chloroplast proteins in Ostreopsis cf. ovata Fukuyo, and Coolia monotis Meunier, which produce different suites of polyketide toxins, suggesting that these PKSs may share common pathways. Since PKSs are closely related to fatty acid synthases (FAS), we sought to determine if fatty acid biosynthesis colocalizes with either chloroplast or cytosolic PKSs. [(3) H]acetate labeling showed fatty acids are synthesized in the cytosol, with little incorporation in chloroplasts, consistent with a Type I FAS system. However, although 29 sequences in a K. brevis expressed sequence tag database have similarity (BLASTx e-value <10(-10) ) to PKSs, no transcripts for either Type I (cytosolic) or Type II (chloroplast) FAS are present. Further characterization of the FAS complexes may help to elucidate the functions of the PKS enzymes identified in dinoflagellates.
authors.affiliation: Marine Biotoxins Program, NOAA Center for Coastal Environmental Health and Biomolecular Research, Charleston, South Carolina, 29412, USA.|Marine Biomedical and Environmental Sciences, Medical University of South Carolina, Charleston, South Carolina, 29412, USA.
authors.forename: Frances M
authors.initials: FM
authors.lastname: Van Dolah
authors.affiliation: Marine Biotoxins Program, NOAA Center for Coastal Environmental Health and Biomolecular Research, Charleston, South Carolina, 29412, USA.|Marine Biomedical and Environmental Sciences, Medical University of South Carolina, Charleston, South Carolina, 29412, USA.
authors.forename: Mackenzie L
authors.initials: ML
authors.lastname: Zippay
authors.affiliation: Interdepartmental Research Centre for Environmental Science (CIRSA), University of Bologna, Ravenna, 48123, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Pezzolesi
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, Florida, 33199, USA.
authors.forename: Kathleen S
authors.initials: KS
authors.lastname: Rein
authors.affiliation: Marine Biotoxins Program, NOAA Center for Coastal Environmental Health and Biomolecular Research, Charleston, South Carolina, 29412, USA.|Marine Biomedical and Environmental Sciences, Medical University of South Carolina, Charleston, South Carolina, 29412, USA.
authors.forename: Jillian G
authors.initials: JG
authors.lastname: Johnson
authors.affiliation: Marine Biotoxins Program, NOAA Center for Coastal Environmental Health and Biomolecular Research, Charleston, South Carolina, 29412, USA.
authors.forename: Jeanine S
authors.initials: JS
authors.lastname: Morey
authors.affiliation: Marine Biotoxins Program, NOAA Center for Coastal Environmental Health and Biomolecular Research, Charleston, South Carolina, 29412, USA.
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Interdepartmental Research Centre for Environmental Science (CIRSA), University of Bologna, Ravenna, 48123, Italy.
authors.forename: Rossella
authors.initials: R
authors.lastname: Pistocchi
keywords: Coolia monotis; Karenia brevis; Ostreopsis cf. ovata; brevetoxin; dinoflagellates; fatty acid synthase; palytoxin; polyketide synthase