Abstract Detail

Comparative Genomics/Transcriptomics

Picard, Colette [1], Povilus, Rebecca [2], Williams, Ben [3], Gehring, Mary [3].

Single nucleus analysis of Arabidopsis endosperm reveals new, transcriptionally distinct cell types.

Endosperm is a biparental seed tissue that is unique to flowering plants and is central to seed development and provisioning. Morphological studies have traditionally defined three main endosperm zones: micropylar, peripheral, and chalazal. New technologies, such as single-nuclei RNAseq, now allow us to define endosperm domains based on transcriptional profiles while simultaneously characterizing distinct suites of gene activity among these domains. We therefore performed RNA sequencing on 1,437 single endosperm nuclei collected from young seeds of crosses involving two genotypes of Arabidopsis thaliana.
We used a clustering strategy to identify up to 14 distinct types of endosperm nuclei, depending on the parental genotypes. Using a set of previously defined marker genes for micropylar, peripheral, and chalazal endosperm tissue, as well as cell-cycle markers, and we assigned initial identity estimates to all endosperm nuclei clusters. We verified these identity assignments by selecting a small number of marker genes for in situ hybridization, to characterize cluster-specific spatial expression patterns within seeds.
Intriguingly, we noticed that two endosperm nuclei clusters were consistently identified as ‘chalazal endosperm’ in all cross types. We performed in situ hybridization for marker genes that were uniquely and highly expressed in these nuclei clusters and discovered that the two ‘chalazal endosperm’ clusters displayed distinct spatial expression domains within the morphologically defined chalazal endosperm zone. Thus, chalazal endosperm consists of two spatially and transcriptionally distinct cell nuclei types: nodules and cysts.
We further characterized the transcriptional profiles of these two types of chalazal endosperm nuclei. The sets of genes significantly up- or down-regulated in the cyst and nodule clusters are enriched in distinct suites of GO terms, suggesting these cell types perform distinct functions. Analysis of cell-cycle markers suggests two thirds of nodule nuclei were in S- or M-phase, while most cyst nuclei were in G1 or G2. This suggests that a major difference between the nodules and cyst is participation in the cell cycle, with chalazal cyst representing the more developmentally fixed, non-dividing tissue.
Understanding the developmental complexity of endosperm is important for understanding how endosperm functions as a mediator between the embryo and maternal seed tissues. Our transcriptionally-based classification of endosperm nuclei represents an important step in characterizing the full complexity of endosperm cell types in Arabidopsis thaliana.

1 - Massachusetts Institute of Technology, Computational and Systems Biology , Cambridge, MA, 02139, USA
2 - Whitehead Institute, Gehring Lab, Whitehead Institute, 455 Main Street, Cambridge, MA, 02142, United States
3 - Whitehead Institute for Biomedical Research, Cambridge, MA, 02142, USA

Single-nuclei sequencing
Seed Biology

Presentation Type: Oral Paper
Number: CG3002
Abstract ID:606
Candidate for Awards:Margaret Menzel Award

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