Maternal and paternal genomes contribute equally to the transcriptome of early plant embryos

Abstract

In animals, maternal gene products deposited into eggs regulate embryonic development before activation of the zygotic genome. In plants, an analogous period of prolonged maternal control over embryogenesis is thought to occur based on some gene-expression studies. However, other gene-expression studies and genetic analyses show that some transcripts must derive from the early zygotic genome, implying that the prevailing model does not fully explain the nature of zygotic genome activation in plants. To determine the maternal, paternal and zygotic contributions to the early embryonic transcriptome, we sequenced the transcripts of hybrid embryos from crosses between two polymorphic inbred lines of Arabidopsis thaliana and used single-nucleotide polymorphisms diagnostic of each parental line to quantify parental contributions. Although some transcripts seemed to be either inherited from primarily one parent or transcribed from imprinted loci, the vast majority of transcripts were produced in near-equal amounts from both maternal and paternal alleles, even during the initial stages of embryogenesis. Results of reporter experiments and analyses of transcripts from genes that are not expressed in sperm and egg indicate early and widespread zygotic transcription. Thus, in contrast to early animal embryogenesis, early plant embryogenesis is mostly under zygotic control.

Figure 1. Maternal and paternal genomes contribute equally to the early embryonic transcriptome

a, Proportion of RNA-Seq reads overlapping maternal and paternal SNPs within transcribed regions of annotated genes. Drawings illustrate stages of isolated embryos, colored to represent the diversity of cell lineages in each stage. Pie charts show proportions of reads overlapping maternal and paternal SNPs. b, Distributions of maternal-to-paternal ratios (log₂) in 1-cell/2-cell (top), 8-cell (middle) and ~32-cell (bottom) embryos. Maternal-to-paternal ratios were calculated for transcripts that had ≥5 SNP-overlapping reads from the Col-0 × Cvi-0 crosses (green; n > 7,388 for each stage) or the Cvi-0 × Col-0 crosses (purple; n > 8,071 for each stage), and transcripts that had ≥5 SNP-overlapping reads from both reciprocal crosses (black; n > 6,892 for each stage). Binomial modeling determined the distributions expected for transcripts examined in both reciprocal crosses if there had been no parent-of-origin effects and no noise apart from stochastic counting variability (gray dashed lines).

Figure 2. Maternally and paternally inherited transgenes are transcribed in the initial stages of embryogenesis

Confocal scanning laser microscopy images show early embryos from the indicated crosses, which were between plants with a transgenic reporter (pOp::GFP) and either plants without an activator (wild-type) or plants with a transgenic activator driven from either the RPS5A or the UBI3 promoter (pRPS5A::LhG4 or pUBI3::LhG4, respectively). Embryonic stages and the approximate number of hours after fertilization (HAF) are shown to the left. GFP signal is indicated by green nuclei, and background autofluorescence is shown in red. Embryos are outlined in white for clarity. Asterisks indicate endosperm nuclei. Of the seeds with GFP signal in endosperm (n in parentheses), the percentage that also had GFP signal in embryos is indicated below each image. In endosperm, expression from maternal alleles is expected to be two-fold higher than that from paternal alleles, but this difference is not observed in these images because the GFP signal in the endosperm was beyond its dynamic range. Scale bars represent 10 µm.

Figure 3. Dozens of transcripts are preferentially inherited or preferentially expressed

a, Maternal-to-paternal ratios for transcripts in 1-cell/2-cell, 8-cell and ~32-cell embryos. Ratios for transcripts that had ≥8 SNP-overlapping reads in both the Col-0 × Cvi-0 and Cvi-0 × Col-0 crosses are plotted. Points, as well as the numbers, that passed cutoffs for maternally or paternally enriched transcripts (see text) are indicated in red and blue, respectively. b, Heat maps of both maternal-to-paternal ratios and changes in transcript levels for transcripts passing cutoffs as maternally (top) or paternally (bottom) enriched in at least one of the three stages. Maternal-to-paternal ratios are color-coded (key), unless the transcript had < 8 SNP-overlapping reads in one cross, in which case the corresponding heat-map value is not colored. Changes in the levels of a transcript (RPM) relative to its mean are color-coded (key). Numbers to the left indicate candidates for both preferential inheritance and preferential expression (1), preferential inheritance (2) or preferential expression (3). Numbers to the right highlight a subset of the class 2 transcripts, which had relatively high levels in 1-cell/2-cell embryos compared to ~32-cell embryos and thus potentially derived mainly from preferential inheritance (2A), or a subset of class 3 transcripts, which had relatively low levels in 1-cell/2-cell embryos compared to ~32-cell embryos and thus potentially derived mainly from parent-of-origin–specific transcription (3A).