No abstract

SUMMARYThe formation of a zygote by the fusion of egg and sperm involves the two gametic transcriptomes. In flowering plants, the embryo sac embedded within the ovule contains the egg cell, whereas the pollen grain contains two sperm cells inside a supporting vegetative cell. The difficulties of collecting isolated gametes and consequent low recovery of RNA have restricted in-depth analysis of gametic transcriptomes in flowering plants. We isolated living egg cells, sperm cells and pollen vegetative cells from Oryza sativa (rice), and identified transcripts for approximately 36 000 genes by deep sequencing. The three transcriptomes are highly divergent, with about three-quarters of those genes differentially expressed in the different cell types. Distinctive expression profiles were observed for genes involved in chromatin conformation, including an unexpected expression in the sperm cell of genes associated with active chromatin. Furthermore, both the sperm cell and the pollen vegetative cell were deficient in expression of key RNAi components. Differences in gene expression were also observed for genes for hormonal signaling and cell cycle regulation. The egg cell and sperm cell transcriptomes reveal major differences in gene expression to be resolved in the zygote, including pathways affecting chromatin configuration, hormones and cell cycle. The sex-specific differences in the expression of RNAi components suggest that epigenetic silencing in the zygote might act predominantly through female-dependent pathways. More generally, this study provides a detailed gene expression landscape for flowering plant gametes, enabling the identification of specific gametic functions, and their contributions to zygote and seed development.

The zygotic transition, from a fertilized egg to an embryo, is central to animal and plant reproduction. Animal embryos depend upon maternally provided factors until zygotic genome activation (ZGA). In plants, the timing and parental genome contributions to ZGA are unresolved. Here, we use the flowering plant Oryza sativa (rice) to characterize transcriptomes of time-staged isogenic and hybrid zygotes following fertilization. Large-scale transcriptomic changes were observed in unicellular zygotes, including upregulation of S-phase genes, a characteristic of ZGA. The parental contributions to ZGA were highly asymmetric. Zygotic transcription was primarily from the maternal genome and included genes for basic cellular processes. Transcription of the paternal genome was highly restricted but unexpectedly included genes encoding putative pluripotency factors expressed at the onset of ZGA. Thus, distinct transcriptional activities are exhibited by the parental genomes during the initiation of embryogenesis, which presumptively derive from divergent pre-zygotic transcriptional states established in the gametes.

The maize (Zea mays) transposable element Dissociation (Ds) was mobilized for large-scale genome mutagenesis and to study its endogenous biology. Starting from a single donor locus on chromosome 10, over 1500 elements were distributed throughout the genome and positioned on the maize physical map. Genetic strategies to enrich for both local and unlinked insertions were used to distribute Ds insertions. Global, regional, and local insertion site trends were examined. We show that Ds transposed to both linked and unlinked sites and displayed a nonuniform distribution on the genetic map around the donor r1-sc:m3 locus. Comparison of Ds and Mutator insertions reveals distinct target preferences, which provide functional complementarity of the two elements for gene tagging in maize. In particular, Ds displays a stronger preference for insertions within exons and introns, whereas Mutator insertions are more enriched in promoters and 59-untranslated regions. Ds has no strong target site consensus sequence, but we identified properties of the DNA molecule inherent to its local structure that may influence Ds target site selection. We discuss the utility of Ds for forward and reverse genetics in maize and provide evidence that genes within a 2-to 3-centimorgan region flanking Ds insertions will serve as optimal targets for regional mutagenesis.