权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Elucidating the Biosynthesis of a Model Ladder-Frame Polyether Toxin

阐明梯架聚醚毒素模型的生物合成

基本信息

批准号：
10066290
负责人：
Timothy R Fallon
金额：
$ 6.49万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-15 至 2023-09-14
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10066290
关键词：
Algal Blooms Anabolism Aquaculture Back Biochemical Biochemistry Biological Models CRISPR screen Candidate Disease Gene Carbon Cataloging Catalogs Cells Cessation of life Chemicals Ciguatoxins Clone Cells Complex Cryopreservation DNA Data Set Development Diatoms Dinophyceae Ecosystem Environment Enzymatic Biochemistry Enzymes Event Experimental Genetics Family Fishes Florida Fractionation Fresh Water Gene Proteins Genes Genetic Genome Genomics Geographic state Gymnodinium breve toxin Haploidy In Vitro Knowledge Link Logic Mammals Methods Modeling Monitor Mutate Natural Products Organism Pathway interactions Production Proteins Public Health Reporting Reproducibility Research Ribonucleoproteins Rice Route Shellfish Source Tandem Repeat Sequences Testing Toxin Transcriptional Regulation Water Supply alpha Toxin base bioaccumulation brevetoxin candidate identification causal variant chlorination contaminated drinking water glycosylation glycosyltransferase harmful algal blooms in vitro testing member metabolomics mutant particle polyketide synthase prevent reconstitution red tide tool transcriptomics

项目摘要

Project Summary/Abstract The societal damage from Harmful Algal Blooms, or HABs, continues to increase globally, with resulting impacts such as fish kills in the wild and in aquaculture, death of marine mammals, and even direct public health concerns though contamination of drinking water supplies or bioaccumulation of HAB toxins in otherwise edible shellfish. DNA based monitoring of HAB toxin biosynthetic genes currently provides a reliable and species-agnostic method to predict the development of a toxic HAB versus an innocuous algal bloom, but the biosynthetic genes for a number of highly impactful HAB compounds are unknown, preventing this monitoring approach across all toxin classes. The large ladder-frame polyether toxins represent one such long standing biosynthetic question, where their long recognized biosynthetic source, namely being polyketide derived natural products, has not yet led to the identification of the causal genes responsible for toxin biosynthesis in any eukaryotic algal producer, such as the “red-tide” dinoflagellates that cause annual toxic “red tide” events in Florida. The lack of clarity regarding dinoflagellate ladder-frame polyether biosynthesis is possibly due to their remarkably intractable genetics. Toxic dinoflagellate species have very large genomes in the 100s of gigabases, genes arrayed in tandem repeats, and a lack of transcriptional regulation, making the causal determinations of gene-chemotype links experimentally difficult. Here, I propose that an alternative toxic microalgae, the haptophyte Prymnesium parvum, producer of the ‘prymnesin’ ladder-frame polyether toxins, and an impactful HAB organism in its own right, is an ideal model system to elucidate the full biosynthetic pathway of a ladder-frame polyether. This proposal aims to identify and characterize the genes and enzymes responsible for prymnesin biosynthesis in Prymnesium parvum. I propose to use computational genomic, transcriptomic, and metabolomic approaches combined with experimental genetic and biochemical approaches to elucidate the biosynthetic pathway for the toxic ‘prymnesin’ ladder-frame polyethers from haploid strains of the experimentally well-suited haptophyte Prymnesium parvum. First, a substrain of P. parvum will be cloned and reference datasets produced and candidate biosynthetic genes cataloged. Second, activity guided fractionation will be used to identify those enzymes in prymnesin biosynthesis using substrates that can be tractably isolated from Prymnesium cultures. Lastly, a functional CRISPR/Cas9 screen will be used to establish causal links to prymnesin biosynthesis for those genes intractable to heterologous reconstitution and in vitro biochemistry. The research would result in a biosynthetic model of prymnesin production, which could be used to develop biosynthetic gene-based monitoring approaches for toxic polyethers in marine and freshwater ecosystems.

项目总结/文摘