Two imprinted genes primed by DEMETER in the central cell and activated by WRKY10 in the endosperm.

The early development of the endosperm is crucial for balancing the allocation of maternal nutrients to offspring. This process is believed to be evolutionarily associated with genomic imprinting, resulting in parentally biased allelic gene expression. Beyond FertilizationIndependentSeed (FIS) genes, the number of imprinted genes involved in early endosperm development and seed size determination remains limited. This study introduces early endosperm-expressed HAIKU (IKU) downstream Candidate F-box 1 (ICF1) and ICF2 as maternally expressed imprinted genes (MEGs) in Arabidopsis thaliana. Although these genes are also demethylated by DEMETER (DME) in the central cell, their activation differs from the direct DME-mediated activation seen in classical MEGs such as the FIS genes. Instead, ICF maternal alleles carry pre-established hypomethylation in their promoters, priming them for activation by the WRKY10 transcription factor in the endosperm. On the contrary, paternal alleles are predominantly suppressed by CG methylation. Furthermore, we find that ICF genes partially contribute to the small seed size observed in iku mutants. Our discovery reveals a two-step regulatory mechanism that highlights the important role of conventional transcription factors in the activation of imprinted genes, which was previously not fully recognized. Therefore, the mechanism provides a new dimension to understand the transcriptional regulation of imprinting in plant reproduction and development.

A simple and highly efficient strategy to induce both paternal and maternal haploids through temperature manipulation.

Haploid production by outcrossing with inducers is one of the key technologies to revolutionize breeding. A promising approach for developing haploid inducers is by manipulating centromere-specific histone H3 (CENH3/CENPA)1. GFP-tailswap, a CENH3-based inducer, induces paternal haploids at around 30% and maternal haploids at around 5% (ref. 2). However, male sterility of GFP-tailswap makes high-demand maternal haploid induction more challenging. Our study describes a simple and highly effective method for improving both directions of haploid production. Lower temperatures dramatically enhance pollen vigour but reduce haploid induction efficiency, while higher temperatures act oppositely. Importantly, the effects of temperatures on pollen vigour and on haploid induction efficiency are independent. These features enable us to easily induce maternal haploids at around 24.8% by using pollen of inducers grown at lower temperatures to pollinate target plants, followed by switching to high temperatures for haploid induction. Moreover, paternal haploid induction can be simplified and enhanced by growing the inducer at higher temperatures pre- and post-pollination. Our findings provide new clues for developing and using CENH3-based haploid inducers in crops.

Cross Inhibition of MPK10 and WRKY10 Participating in the Growth of Endosperm in Arabidopsis thaliana.

The product of double fertilization produces seed, which contains three components: triploid endosperm, diploid embryo, and maternal seed coat. Amongst them, the endosperm plays a crucial role in coordinating seed growth. Mitogen-activated protein kinase (MAPK) cascades are conserved in eukaryotes and involved in signal transduction of plant development. MPK3, MPK6, and MPK10 form a small group of MPKs family in Arabidopsis thaliana. MPK3 and MPK6 are extensively studied and were found to be involved in diverse processes including plant reproduction. However, less is known about the function of MPK10. Here, we found WRKY10/MINI3, a member of HAIKU (IKU) pathway engaging in endosperm development, and MPK10 is high-specifically expressed in the early developmental endosperm but with opposite gradients. We further proved that MPK10 and WRKY10 cross-inhibit the expression of each other. The inhibition effect of MPK10 on gene expression of WRKY10 and the downstream targets is supported by the fact that MPK10 interacts with WRKY10 and suppresses the transcriptional activity of WRKY10. Constantly, mpk10 mutants produce big seeds while WRKY10/MINI3 positively regulate seed growth. Altogether, our data provides a model of WRKY10 and MPK10 regulating endosperm development with a unique cross inhibitory mechanism.