Research Article

RETINOBLASTOMA RELATED1 mediates germline entry in Arabidopsis

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Science  28 Apr 2017:
Vol. 356, Issue 6336, eaaf6532
DOI: 10.1126/science.aaf6532

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Germ cells on demand

Unlike animals, plants do not set aside a germline. Instead, germ cells are developed on demand from somatic lineages. Zhao et al. examined the regulatory pathways that manage the transition from somatic to germ cell development in the small plant Arabidopsis (see the Perspective by Vielle-Calzada). The transcription factor WUSCHEL (WUS) was needed early on for development of ovules. Soon after, a trio of inhibitors that work through a cyclin-dependent kinase allowed a transcriptional repressor to down-regulate WUS. This opened the door to meiosis, while restricting the number of reproductive units per seed to one.

Science, this issue p. eaaf6532; see also p. 378

Structured Abstract

INTRODUCTION

Seeds of flowering plants have evolved to typically carry only a single embryo next to a nourishing tissue, the endosperm that functions analogously to the human placenta. Both structures are formed from a single female gametophyte (embryo sac) that develops from one meiotic product, the functional megaspore, in most sexually reproducing plants. As in many other organisms, including humans, all but one female meiotic product die to assure the development of only one reproductive unit per future seed.

RATIONALE

In contrast to humans and animals, plants do not set aside a specialized cell lineage (germline) that produces meiocytes in early embryogenesis. Instead, the germline of plants is established de novo from somatic cells in floral reproductive organs. Several genes have been identified that control the formation of ovules, which harbor the meiocytes [megaspore mother cells (MMCs)]. These include the homeodomain transcription factor WUSCHEL (WUS), a key regulator of stem cell fate in plants that is essential for the formation of the integuments from which the seed coat is derived. Moreover, WUS is also involved in the specification of MMCs. However, it is not clear how somatic cells that divide mitotically switch to a meiotic cell division program.

RESULTS

Following up expression data of young ovules primordia and MMCs, we reveal a regulatory cascade that controls the entry into meiosis, starting with a group of redundantly acting cyclin-dependent kinase (CDK) inhibitors of the KIP-RELATED PROTEIN (KRP) class. KRPs function by restricting CDKA;1–dependent inactivation of the Arabidopsis Retinoblastoma homolog RBR1. In rbr1 and krp triple mutants, designated meiocytes undergo several mitotic divisions, resulting in the formation of supernumerary meiocytes that give rise to multiple reproductive units per future seed. Live observation revealed that these multiple units can successfully attract a pollen tube and can be fertilized. However, subsequent seed development is blocked, resulting in semisterility of the mutant plants. One of the functions of RBR1 is the direct repression of the stem cell factor WUSCHEL (WUS), which ectopically accumulates in meiocytes of triple krp and rbr1 mutants. Depleting WUS in rbr1 mutants restored the formation of only a single meiocyte. However, ectopic expression of WUS by itself is not sufficient to induce mitotic divisions instead of meiosis, revealing that RBR1 is a central hub of meiocyte differentiation.

CONCLUSION

There is a delicate balance between WUS activation important for ovule primordia formation—including the development of the integuments, as well as a role in specifying the MMC itself—and its inactivation by RBR1 soon afterward to allow entry into meiosis. Different components of the Rb control pathway have been associated with germ cell fate initiation in animals; for example, mutants in the CDK inhibitor dacapo in Drosophila fail to enter meiosis. Similarly, down-regulation of Cdk2-cyclin E, a well-known regulator of Rb, is important for Caenorhabditis elegans germline development. This raises the intriguing question of whether Rb family proteins play a conserved role in germline entry in multicellular organisms.

Formation of multiple MMCs in Arabidopsis.

The left column shows the wild type, the middle column shows krp4 krp6 krp7 triple mutants, and the right column shows rbr1-2. A designated MMC undergoes a mitotic instead of a meiotic division, leading to the production of multiple MMCs in triple krp and rbr1 mutants (top row) instead of a single MMC, as found in the wild type. MMC fate is highlighted by means of KNU-YFP expression (second row). Multiple MMCs give rise to multiple gametophytes in ovules (third row and highlighted in different shades of blue in the bottom row).

Abstract

To produce seeds, flowering plants need to specify somatic cells to undergo meiosis. Here, we reveal a regulatory cascade that controls the entry into meiosis starting with a group of redundantly acting cyclin-dependent kinase (CDK) inhibitors of the KIP-RELATED PROTEIN (KRP) class. KRPs function by restricting CDKA;1–dependent inactivation of the Arabidopsis Retinoblastoma homolog RBR1. In rbr1 and krp triple mutants, designated meiocytes undergo several mitotic divisions, resulting in the formation of supernumerary meiocytes that give rise to multiple reproductive units per future seed. One function of RBR1 is the direct repression of the stem cell factor WUSCHEL (WUS), which ectopically accumulates in meiocytes of triple krp and rbr1 mutants. Depleting WUS in rbr1 mutants restored the formation of only a single meiocyte.

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