Will it Frankenstein? Discovering the role of immune-based rejection in incompatible plant grafts

When you pick a flower, easily breaking its stem, it’s hard to imagine how resilient this organism is. Plants can survive so much, and are capable of regenerating leaves, branches, or even whole shoots from the root when damaged. Amazingly, some plants can even merge themselves together with another plant, creating a whole new plant. This technique is known as grafting, where the apical portion of one plant, known as the scion, is connected to the root system, known as the rootstock, of another plant. Since the beginning of agriculture, the hunt has been on to create higher crop yields that will feed Earth’s continuously growing population. While current agricultural technologies such as conventional breeding of more resistant crops or creating transgenic plants can introduce resistance to abiotic and biotic stresses, including disease and cold tolerance, they take years to develop and be approved. Grafting provides these same benefits, but at a more efficient rate than breeding. In order for grafting to be successful, the species must be able to reform vascular connection. When the scion and rootstock do not form vascular connections with each other, the two species are considered graft incompatible. However, there is little known about the molecular basis of graft incompatibility. This is a significant issue for farmers, who utilize grafting to increase their agricultural production. In molecular studies of incompatible grafting, tomato and pepper grafts are often used as a model system. Tomato and pepper graft easily to themselves, while tomato:pepper grafts are known to be incompatible. Interestingly, recent studies in our lab of incompatible tomato:pepper grafts have found over 1000 upregulated genes that are also involved in immune response, cell death, and other defense-related processes, indicating a possible immune response in incompatible grafts. In this study, we seek to determine whether cross-species graft incompatibility is caused by immune response. By using tomato mutants with knockouts of known disease regulatory pathways, we can study the effect of loss of function in disease pathways on graft compatibility in normally compatible tomato:tomato self-grafts, as well as in incompatible tomato:pepper grafts. Through characterization of mutant graft phenotypes with a novel Toluidine blue flash-staining method for visualizing vascular connectivity, we can determine the functional role of disease regulatory networks in graft compatibility. We observed a change in graft compatibility in disease mutant lines, providing experimental evidence of disease response in graft compatibility.

I am incredibly grateful to have had the opportunity to work in the Frank lab through the Boyce Thompson Institute and Cornell University’s Plant Genome Research Program REU. Coming from a university with limited plant science opportunities, I knew I wanted to gain more experience in plant biology and explore the different career options the field offers. This experience gave me everything I could have imagined and more. In the Frank lab, I was co-mentored by Sam Yanders and Margaret Frank and worked on three projects. Each of these projects taught me how to manage my time effectively and become a more independent researcher, giving me unique experiences that have shown me what it is like to be a PhD student.

The first project I worked on was characterizing ROS and calcium reporter lines in different tomato varieties, which helped me refine my genotyping skills. The second project was graft characterization of disease mutants in tomato self grafts and tomato : pepper heterografts. This project introduced me to grafting and gave me the opportunity to create my own rapid and robust staining protocol. The third project I worked on was generating agrobacterium symbionts in tomato : pepper heterografts to see if these symbionts could rescue failed vascular connections in incompatible grafts. This taught me how to design my own project and think about the quantity and type of variables I wanted to test. Not only did I gain a multitude of lab techniques and experiences, but I also have memories that I will remember for a lifetime. One of these memories is sitting next to one of my mentors, Sam, and having her trying out the staining protocol I created on disease mutant self grafts. Not only was she able to get the protocol to work, but we also found that the mutant self graft had a significantly different phenotype compared to the wild type self graft, revealing that immunity based responses play a role in grafting compatibility and that this project will now be her thesis defense!

Outside of the lab, I found an amazing group of friends and gained experience in science communication, learning how I can bridge the gap between research and the public. Participating in the REU program has given me the confidence to pursue a PhD in plant biology and how I can use my research to connect with the public and make a greater mpact on the world.