RUI: Understanding the cellular basis of seed filling in the late Arabidopsis thaliana embryo

For thousands of years seeds have been a crucial part of our diet and that of domesticated animals. Seeds are nutritious because during the latter part of their development (maturation) they accumulate starch, protein and fats (seed filling), to feed the germinating seedling. The regulation of the seed filling process is only partially understood.

In particular, what controls the dynamic way in which seed filling genes are turned on or off only in certain cells and at certain times within embryos inside the seeds is unclear. It is crucial, then, to study gene expression at the single-cell level, which will be done using a novel technique called single-nucleus RNA sequencing. What this means is to isolate plant embryos of the right developmental stages, purify their nuclei, and see which genes are on (“expressed”) in each cell.

The process will be studied in the model plant Arabidopsis thaliana, of the mustard and canola family. These data will provide a three-dimensional atlas that indicates which genes are expressed when and where. Computational techniques will then be applied to this dataset to help us reconstruct “gene regulatory networks”, that is, diagrams that show which genes control which others.

Using these networks, new genes that control maturation and seed filling are expected to be found. This atlas will be useful for other members of the research community studying seed maturation and could contribute to improving nutritional content in seed crops. Additionally, this project will provide undergraduate students with hands-on experience in cutting-edge research, helping to advance the nation’s scientific competitiveness.

Outreach will be performed through a weekly science show on a public radio station, and through activities with a local high school.

The overall goal of this project is to increase the understanding of the maturation phase in dicot embryos and the regulation of the accompanying accumulation of storage products (seed filling). The onset of the maturation program dramatically changes the gene expression profile of the seed and is tightly regulated. The seed filling process is a spatially dynamic process, starting in certain tissues and later expanding to the rest of the embryo.

The spatial aspects of this developmental transition have never been carefully examined, and it is unclear how the tissue-specific expression of the genes involved is controlled. Prior transcriptomic analyses of maturation phase embryos have all employed whole embryos. Here, embryos will be sampled from key time points during seed maturation and single-nucleus RNAseq will be used to generate a publicly accessible three dimensional database of cellular states.

Tissue-specific gene regulatory networks will be reconstructed and lead to the expected finding of novel regulators of seed filling, which will later be validated and characterized.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.