Deciphering the eccDNA inheritance with novel plant eccDNA genetic tools

Project Information

circlePLANT

Grant agreement ID: 101068070

DOI

10.3030/101068070

Project closed

EC signature date 3 June 2022

Start date 1 September 2022

End date 31 August 2024

Funded under

• Marie Skłodowska-Curie Actions (MSCA)

Total cost

No data

EU contribution

€ 214 934,40

Coordinated by

KOBENHAVNS UNIVERSITET
Denmark

Periodic Reporting for period 1 - circlePLANT (Deciphering the eccDNA inheritance with novel plant eccDNA genetic tools)

Reporting period: 2022-09-01 to 2024-08-31

Summary of the context and overall objectives of the project

Global challenges such as climate change and food insecurity demand innovative solutions to increase crop resilience and productivity while reducing environmental impact. By 2050, the global population is projected to reach 9.1 billion, requiring a 70% increase in food production. Current agricultural practices rely heavily on fertilizers, herbicides, and pesticides, which have caused substantial soil degradation, water eutrophication, and depletion of non-renewable nutrient sources. In response, the European Union’s Common Agricultural Policy (CAP), European Green Deal, and Farm to Fork strategy emphasize sustainable, low-input agriculture to balance productivity and environmental health.

The circlePLANT project explored the role of extrachromosomal circular DNA (eccDNA) in plants as a potential mechanism for stress adaptation and transgenerational inheritance of stress memory. EccDNA, a unique form of genetic material found in all eukaryotic organisms, may carry stress-responsive genetic information across generations, providing plants with an adaptive advantage under adverse environmental conditions.

The project aimed to address two key objectives:

Develop a genetic toolbox to study eccDNA biogenesis in plants, focusing on CRISPR/Cas9-based systems to create and track specific eccDNAs.

Characterize the inheritance of eccDNAs, particularly their transmission during meiosis from the parental sporophyte to gametes and progeny, with an emphasis on heat stress (HS)-induced eccDNAs.

These objectives were designed to advance fundamental plant biology, inform strategies for improving plant stress resilience, and align with European Union sustainability goals.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The circlePLANT project carried out an extensive investigation into the diversity and dynamics of extrachromosomal circular DNA (eccDNA) in Arabidopsis. A genome-scale analysis identified over half a million unique eccDNAs across different tissues and temperature treatments. This comprehensive mapping effort provided a robust foundation for exploring eccDNA characteristics, offering one of the most detailed catalogs of these elements in plants to date.
The project also investigated the mechanisms behind eccDNA formation, focusing on the role of DNA damage. Experiments demonstrated that double-strand breaks (DSBs) in DNA, induced by treatments such as radiomimetic agents, are a critical trigger for eccDNA biogenesis. These findings confirmed previous assumptions about the connection between genome instability and eccDNA formation, providing valuable insights for future studies on their regulation.
Additionally, the analysis of recurrent eccDNAs revealed that they contain tandem duplications, implicating homologous recombination (HR) as a key mechanism in their formation. This result supports the hypothesis that homologous regions play a central role in driving eccDNA generation, offering a mechanistic understanding of their origins.
To facilitate further research, the project developed high-quality datasets and two dedicated databases: one cataloging the diversity of eccDNAs across Arabidopsis tissues and treatments and another focusing on quantitative metrics of gene circularization. These resources, designed to be openly accessible, provide the scientific community with tools to study eccDNA dynamics and functions systematically.
Despite technical challenges with CRISPR/Cas9-based methods, the project made significant progress through alternative approaches, achieving its goal of delivering foundational insights into the biology of eccDNAs in plants.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The circlePLANT project uncovered groundbreaking findings that significantly advance the understanding of eccDNA in plant stress adaptation and inheritance. Previous research on eccDNA in plants was limited to their presence and association with repetitive DNA. This project went further, delivering the first genome-scale analysis of eccDNAs, uncovering their functional roles, and revealing their potential as mechanisms for transmitting adaptive information.
One of the project’s most novel discoveries was the persistence of heat-stress-induced gene circularization patterns across tissues and generations. These patterns, first observed in plants exposed to heat stress during early sporophyte development, were retained in tissues that developed later under normal conditions and were also present in the gametophyte and progeny. This suggests that gene circularization may serve as a potential novel mechanism for transmitting stress-response information across life cycle stages, contributing to molecular memory in plants.
The functional relevance of these circularized genes was revealed through their enrichment in biological processes essential for abiotic stress adaptation, such as water regulation, heat response, and protective compound synthesis. These findings go beyond merely cataloging eccDNAs, positioning gene circularization as a potential driver of plant resilience to environmental challenges.
The project also demonstrated that certain eccDNAs, including 89 shared between sporophyte, gametophyte, and progeny, may reflect a mode of inheritance or recurrent formation. This result highlights the dynamic nature of eccDNAs, suggesting they play a role in genomic responses across generations.
By identifying the processes driving eccDNA formation and revealing their implications for stress adaptation and inheritance, the project has laid the groundwork for future applications. These include exploring gene circularization as a tool for developing stress-resilient crops and furthering the understanding of genome plasticity in response to environmental challenges.
The circlePLANT findings align with European policy goals, such as the Green Deal and Farm to Fork Strategy, by contributing knowledge critical to sustainable agriculture and climate resilience. The novel insights into eccDNA inheritance and stress adaptation represent a major step forward, offering exciting opportunities for future research and agricultural innovation.