Award Abstract # 1645256
IOS EDGE: Development of genetic and genomic resources for milkweed, Asclepias syriaca and Asclepias curassavica

NSF Org: IOS
Division Of Integrative Organismal Systems
Recipient: BOYCE THOMPSON INSTITUTE FOR PLANT RESEARCH INC
Initial Amendment Date: March 7, 2017
Latest Amendment Date: March 7, 2017
Award Number: 1645256
Award Instrument: Standard Grant
Program Manager: Diane Jofuku Okamuro
dokamuro@nsf.gov
 (703)292-4508
IOS
 Division Of Integrative Organismal Systems
BIO
 Directorate for Biological Sciences
Start Date: March 15, 2017
End Date: February 28, 2021 (Estimated)
Total Intended Award Amount: $1,020,000.00
Total Awarded Amount to Date: $1,020,000.00
Funds Obligated to Date: FY 2017 = $1,020,000.00
History of Investigator:
  • Georg Jander (Principal Investigator)
    gj32@cornell.edu
  • Steven Broyles (Co-Principal Investigator)
  • Anurag Agrawal (Co-Principal Investigator)
  • Lukas Mueller (Co-Principal Investigator)
Recipient Sponsored Research Office: Boyce Thompson Institute Plant Research
533 TOWER RD
ITHACA
NY  US  14853-7202
(607)254-1248
Sponsor Congressional District: 19
Primary Place of Performance: Boyce Thompson Institute Plant Research
533 Tower Road
Ithaca
NY  US  14853-1801
Primary Place of Performance
Congressional District:
19
Unique Entity Identifier (UEI): ULBJY4YK9ZX8
Parent UEI:
NSF Program(s): Physiol Mechs & Biomechanics
Primary Program Source: 01001718DB NSF RESEARCH & RELATED ACTIVIT
Program Reference Code(s): 050Z, 9178
Program Element Code(s): 765800
Award Agency Code: 4900
Fund Agency Code: 4900
Assistance Listing Number(s): 47.074

ABSTRACT

Milkweed plants are well-known for their iconic interactions with monarch butterflies. Although milkweeds produce toxins that are lethal to most insects, monarch butterfly caterpillars are quite resistant and even store milkweed toxins for their own defense against predators like birds. As is indicated by the milkweed genus name, Asclepias, which is derived from Asclepius, the Greek god of healing, Native Americans used these plants to treat a variety of medical ailments. Several studies have shown that milkweeds are rich sources of novel chemical compounds that not only provide defense against insects, but also have potential medical applications. In this project the investigators will develop new resources to investigate the function of specific genes in both common milkweed and tropical milkweed. Methods will be developed to knock out or overexpress specific milkweed genes in a targeted manner. Genome sequencing of two milkweed species, together with natural variation in milkweed gene expression and chemical content, will enable the discovery of new genes involved in defense against herbivory and the production of medically relevant plant metabolites. The resources that will be developed through this project will be made publicly available as they are being generated, with no restrictions limiting their use. The project also will involve training high school students and undergraduate summer interns, which will prepare them for future research careers in industry, academia, or government service.

The further development of Asclepias syriaca (common milkweed) and Asclepias curassavica (tropical milkweed) as research model systems will require improved tools for studying the functions of individual genes. In this project, milkweed-specific protocols will be developed for: (i) transient gene expression using Agrobacterium infiltration, (ii) virus-induced silencing of endogenous gene expression, (iii) targeted mutagenesis using CRISPR/Cas9, (iv) stable transformation in tissue culture, (v) regeneration of plants from callus, and (vi) heritable modification of the milkweed genome. To facilitate functional analysis of milkweed genes, the A. syriaca and A. curassavica genomes will be assembled using Pacific Biosciences DNA sequencing. Assembled contigs will be ordered and assigned to specific chromosomes using classical genetic maps and BioNano optical maps. Milkweed genes will be identified and annotated through in silico comparisons and deep sequencing of RNA from different tissue types. Genotyped A. syriaca and A. curassavica genetic mapping populations will enable quantitate trait locus mapping of any phenotypic or molecular trait that varies in the constituent lineages. Plant gene expression and metabolite content varies dramatically in different tissue types and in response to environmental stimuli. In particular, insect attack induces the production of a wide variety of biologically active plant metabolites. Transcriptomic and metabolomic analysis of six A. syriaca and A. curassavica tissue types, with and without herbivory treatment, will provide a broad overview of the genetic and metabolic potential of these species. Molecular protocols, genome sequences, transcriptome data, and correlative analyses linking the transcriptomes, metabolomes, and other milkweed phenotypes will be displayed on the project website, www.milkweedbase.net.

PUBLICATIONS PRODUCED AS A RESULT OF THIS RESEARCH

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(Showing: 1 - 10 of 11)
Agrawal, Anurag A. "Advances in understanding the long-term population decline of monarch butterflies" Proceedings of the National Academy of Sciences , v.116 , 2019 10.1073/pnas.1903409116 Citation Details
Agrawal, Anurag A. and Böröczky, Katalin and Haribal, Meena and Hastings, Amy P. and White, Ronald A. and Jiang, Ren-Wang and Duplais, Christophe "Cardenolides, toxicity, and the costs of sequestration in the coevolutionary interaction between monarchs and milkweeds" Proceedings of the National Academy of Sciences , v.118 , 2021 https://doi.org/10.1073/pnas.2024463118 Citation Details
Boege, Karina and Agrawal, Anurag A and Thaler, Jennifer S "Ontogenetic strategies in insect herbivores and their impact on tri-trophic interactions" Current Opinion in Insect Science , v.32 , 2019 10.1016/j.cois.2018.11.004 Citation Details
Broyles, Steven B. and Elkins, Geordie "Hybridization between Asclepias purpurascens and Asclepias syriaca (Apocynaceae): A cause for concern?1" The Journal of the Torrey Botanical Society , v.146 , 2019 10.3159/TORREY-D-18-00050.1 Citation Details
Broyles, Steven B and Stoj, Kiley R "Patterns of nectar production in Asclepias curassavica (Apocynaceae)" Journal of pollination ecology , v.25 , 2019 Citation Details
Goud, Ellie M. and Sparks, Jed P. and Fishbein, Mark and Agrawal, Anurag A. and Shefferson, ed., Richard "Integrated metabolic strategy: A framework for predicting the evolution of carbonwater tradeoffs within plant clades" Journal of Ecology , v.107 , 2019 https://doi.org/10.1111/1365-2745.13204 Citation Details
Jones, Patricia L. and Petschenka, Georg and Flacht, Lara and Agrawal, Anurag A. "Cardenolide Intake, Sequestration, and Excretion by the Monarch Butterfly along Gradients of Plant Toxicity and Larval Ontogeny" Journal of Chemical Ecology , v.45 , 2019 10.1007/s10886-019-01055-7 Citation Details
Mirzaei, Mahdieh and Züst, Tobias and Younkin, Gordon C. and Hastings, Amy P. and Alani, Martin L. and Agrawal, Anurag A. and Jander, Georg "Less Is More: a Mutation in the Chemical Defense Pathway of Erysimum cheiranthoides (Brassicaceae) Reduces Total Cardenolide Abundance but Increases Resistance to Insect Herbivores" Journal of Chemical Ecology , v.46 , 2020 https://doi.org/10.1007/s10886-020-01225-y Citation Details
Züst, Tobias and Mirzaei, Mahdieh and Jander, Georg "Erysimum cheiranthoides, an ecological research system with potential as a genetic and genomic model for studying cardiac glycoside biosynthesis" Phytochemistry Reviews , v.17 , 2018 10.1007/s11101-018-9562-4 Citation Details
Züst, Tobias and Mou, Sophie and Agrawal, Anurag A. and Biere, ed., Arjen "What doesnt kill you makes you stronger: The burdens and benefits of toxin sequestration in a milkweed aphid" Functional Ecology , v.32 , 2018 10.1111/1365-2435.13144 Citation Details
Züst, Tobias and Strickler, Susan R and Powell, Adrian F and Mabry, Makenzie E and An, Hong and Mirzaei, Mahdieh and York, Thomas and Holland, Cynthia K and Kumar, Pavan and Erb, Matthias and Petschenka, Georg and Gómez, José-María and Perfectti, Francisc "Independent evolution of ancestral and novel defenses in a genus of toxic plants (Erysimum, Brassicaceae)" eLife , v.9 , 2020 https://doi.org/10.7554/eLife.51712 Citation Details
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PROJECT OUTCOMES REPORT

Disclaimer

This Project Outcomes Report for the General Public is displayed verbatim as submitted by the Principal Investigator (PI) for this award. Any opinions, findings, and conclusions or recommendations expressed in this Report are those of the PI and do not necessarily reflect the views of the National Science Foundation; NSF has not approved or endorsed its content.

All known plants accumulate a variety of toxic and deterrent metabolites as protection against herbivory. For instance, several plant families have independently evolved the production of cardiac glycosides, which are toxic to most animals. Insects that are specialized for feeding on cardiac glycoside-producing plants, for example monarch butterflies (Danaus plexippus), typically have specific mutations that make them resistant to the toxic effects. At lower doses, plant-produced cardiac glycosides have uses in human medicine for the treatment of congestive heart failure and other diseases. However, although there is extensive research on the ecological role of cardiac glycosides, as well as centuries of use in both traditional and modern medicine, the biosynthetic pathway is not known from any plant species. To facilitate further research on the biosynthesis, ecological functions, and medical applications of cardiac glycosides, genetic and genomic resources were developed for three cardiac glycoside-producing plant species: common milkweed (Asclepias syriaca), tropical milkweed (Asclepias curassavica), and wormseed wallflower (Erysimum cheiranthoides).

A single natural isolate of common milkweed was chosen for genome sequencing and a high-quality genome assembly has been deposited in public databases. Several hundred additional common milkweed isolates from the eastern United States were collected for measurements of genetic relatedness, gene expression levels, and cardiac glycoside content. Although there is geographic variation in the metabolite content and other plant phenotypes, genomic comparisons showed that common milkweed represents a single panmictic population. Correlative analysis of gene expression levels and cardiac glycoside content in this collection of milkweed isolates will lead to the identification of genes that influence cardiac glycoside abundance and diversity.

In the case of tropical milkweed, a single plant line was inbred for six generations to increase homozygosity prior to genome sequencing. Seeds of this genome-sequenced milkweed line represent a permanent resource that will be available for future research. Fifty additional isolates of tropical milkweed were self-pollinated for at least three generations to generate an inbred population that can be used for genetic studies of natural variation in cardiac glycoside content and other traits.

Similar to the approach that was used for tropical milkweed, a sixth-generation inbred line of wormseed wallflower was subjected to genome sequencing. Seeds of this inbred wallflower line have been made available in public stock centers. Additionally, both cardiac glycoside content and gene expression levels were measured in wormseed wallflower and 47 other wallflower species. A comparative analysis showed high metabolic diversity and relatively recent evolution of cardiac glycoside biosynthesis in the wallflower genus. To facilitate future research, methods were developed for stable transformation and virus-mediated transient gene expression in wormseed wallflower. Candidate genes from other cardiac glycoside-producing plant species can be expressed in wormseed wallflower to investigate their functions.

In the course of this project, graduate students and postdocs received training in plant genetics, genomics, and metabolite analysis. Additionally, students participating in an NSF-funded Research Experience for Undergraduates program conducted research with both milkweeds and wallflowers. The iconic interaction between common milkweed and monarch butterflies lends itself well to education and outreach activities. Demonstration experiments showing this milkweed-monarch butterfly interaction were set up for local grade school students and at a science center outreach event for the general public.

 


Last Modified: 06/28/2021
Modified by: Georg Jander

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