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Successful tree modifications should be gentle and not compromise plant growth and development

Published: 12 November 2021
Evgeniy Donev

Evgeniy Donev, PhD student in Ewa Mellerowicz group at UPSC, investigated different modification strategies to genetically improve hybrid aspen for biofuel production. The idea is to make sugars in wood cell walls that are the basis for biofuel production, better accessible by modifying the cell wall structure. Based on his results, Evgeniy Donev suggests using gentle modifications restricted to certain tissues and test them both in the greenhouse and under field conditions. He also advices to be cautious when introducing fungal proteins because they can trigger an immune response just by its presence. Evgeniy Donev will defend his PhD thesis at the Swedish University of Agricultural Sciences on Tuesday, 16th of November 2021.

How did you get to know about your PhD project and what aroused your interest in it?

Nicolas Delhomme and Nathaniel Street from UPSC were teachers of mine during my master’s degree in Civil Engineering in Biotechnology at Umeå University. Via a student bioinformatic project, I met my PhD supervisor Ewa Mellerowicz. Already during my studies, I realized that I do not only want to do bioinformatics but also know more about the biology behind. Ewa Mellerowicz planned a whole series of experiments and wanted to analyse them with a broad range of different techniques including also complex methods that produce a vast amount of data and require some bioinformatic knowledge. I was very interested in this and when I finished my master thesis in her group, I did not want to finish the project. Luckily, she thought the same and offered me to continue as a PhD student.

You chose as subtitle of your thesis “From design to the field”. Why do you think is this important?  

My thesis addresses several problems important for improving trees - in our case hybrid aspen - for biorefinery and biofuel production. We identified the best engineering strategies based on experiments in the greenhouse and outside in the field. To evaluate the performance of promising genetically improved plants, it is important to test them in an environment which represents their usual cultivation conditions. In many cases, genetically modified plants grow well in the greenhouse but show undesirable reactions in the field conditions where they must cope with a multitude of different stresses like drought or pathogen attacks. Such undesirable reactions or off-target effects can erase all positive effects coming from the gene modification.

What is the best engineering strategy to modify hybrid aspen trees for biofuel production?

We investigated a collection of hybrid aspen in which cell wall properties were modified in different ways to make cellulose and the contained sugars better accessible for biofuel production. Trees in which the modification was introduced to the whole plant developed stronger off-target effects than trees in which the modification was restricted to certain tissues of the plant like for example the wood.

Some of our trees grew better in the greenhouse and in the field. The relative sugar amount that we extracted from those trees was not much higher than in non-modified trees, but they contributed with more biomass. So, the total amount of sugars was higher. On the contrary, we saw that we lose already all wins gained through the modification when the modified trees grew twenty percent less than non-modified trees. Our conclusion is that nothing can replace reduction in growth, and we should therefore choose gentle and more specific engineering strategies. 

Which of your results was the most unexpected for you?

We expressed a protein from a wood decaying fungus in hybrid aspen that is supposed to loosen up the cell wall structure and thus make cellulose better accessible. It looked like the improvement worked because we could extract more sugar from these trees, but the off-target effects were really strong. The trees were dwarf, dropped their leaves very early and showed strong immune defence reactions. Only when we restricted the activity of the protein to the wood, all off-target effects were avoided, and the plants looked normal. However, I started to wonder if just the presence of the fungal gene could be recognised by the plant and trigger an immune response. To test this, we introduced an inactive version of the protein into hybrid aspen, and we saw the same strong effects on the plant. It turned out that the protein we introduced is recognized by the plant as pathogenic and causes an immune response. Sugars are remobilized as part of this response to supply the plant immune system with the necessary energy,  which could possibly affect the sugar concentration extracted from the wood of these plants. 

What were the biggest challenges you faced during your PhD?

My group is focussing on cell wall formation in the wood and not on plant-pathogen interaction. It was very challenging to convince them that the change we saw in those trees in which the fungal protein was active all over the plant, is coming from the reaction of the immune system and not from the activity of the introduced fungal protein. I had to dig deep into the available literature and develop my analytical skills, think through my story thoroughly and keep on discussing with my group. I am very grateful to Ewa Mellerowicz because her support and trust in my skills combined with her positively demanding attitude was motivating me a lot during this time.

What are you planning to do now?

Research in plant science will be in one way or the other part of my future, not least that I see that there are many open questions that are not answered yet. I plan to stay in the group of Ewa Mellerowicz for the next year. We have several very exciting projects which we are currently working on, and I hope that we generate useful knowledge that helps to better understand the complex plant system.

Text: Anne Honsel


About the public defence:

Evgeniy Donev, Umeå Plant Science Centre, Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences, will defend his PhD thesis on Tuesday, 16th of November 2021. Faculty opponent will be Sharon Regan, Department of Biology, Queens University, Kingston Ontario, Canada. The thesis was supervised by Ewa Mellerowicz. The dissertation will be live broadcasted via Zoom.

Title of the thesis: Modification of forest trees by genetic engineering - From design to the field