Summary: Upon hormonal signaling, ovules develop as lateral organs from the placenta. Ovule numbers ultimately determine the number of seeds that develop, and thereby contribute to the final seed yield in crop plants. We demonstrate here that CUP‐SHAPED COTYLEDON 1 (CUC1), CUC2 and AINTEGUMENTA (ANT) have additive effects on ovule primordia formation. We show that expression of the CUC1 and CUC2 genes is required to redundantly regulate expression of PINFORMED1 (PIN1), which in turn is required for ovule primordia formation. Furthermore, our results suggest that the auxin response factor MONOPTEROS (MP/ARF5) may directly bind ANT, CUC1 and CUC2 and promote their transcription. Based on our findings, we propose an integrative model to describe the molecular mechanisms of the early stages of ovule development.

Synchronized communication between gametophytic and sporophytic tissue is crucial for successful reproduction, and hormones seem to have a prominent role in it. Here, we studied the role of the Arabidopsis (Arabidopsis thaliana) cytochrome P450 CYP78A9 enzyme during reproductive development. First, controlled pollination experiments indicate that CYP78A9 responds to fertilization. Second, while CYP78A9 overexpression can uncouple fruit development from fertilization, the cyp78a8 cyp78a9 loss-of-function mutant has reduced seed set due to outer ovule integument development arrest, leading to female sterility. Moreover, CYP78A9 has a specific expression pattern in inner integuments in early steps of ovule development as well as in the funiculus, embryo, and integuments of developing seeds. CYP78A9 overexpression did not change the response to the known hormones involved in flower development and fruit set, and it did not seem to have much effect on the major known hormonal pathways. Furthermore, according to previous predictions, perturbations in the flavonol biosynthesis pathway were detected in cyp78a9, cyp78a8 cyp78a9, and empty siliques (es1-D) mutants. However, it appeared that they do not cause the observed phenotypes. In summary, these results add new insights into the role of CYP78A9 in plant reproduction and present, to our knowledge, the first characterization of metabolite differences between mutants in this gene family.

In spermatophytes the ovules upon fertilization give rise to the seeds. It is essential to understand the mechanisms that control ovule number and development as they ultimately determine the final number of seeds and, thereby, the yield in crop plants. In Arabidopsis thaliana, ovules arise laterally from a meristematic tissue within the carpel referred to as placenta. For a correct determination of the number of ovules, a precise establishment of the positions where ovule primordia emerge is needed, and a tight definition of the boundaries between ovules is therefore also required. In the last decades, few factors have been identified to be involved in the determination of ovule number. Recently, plant hormones have also been revealed as fundamental players in the control of the initiation of ovule formation. In this review we summarize the current knowledge about both the molecular and hormonal mechanisms that control ovule formation in Arabidopsis thaliana.

The developmental programme of the pistil is under the control of both auxin and cytokinin. Crosstalk between these factors converges on regulation of the auxin carrier PIN-FORMED 1 (PIN1). Here, we show that in the triple transcription factor mutant cytokinin response factor 2 (crf2) crf3 crf6 both pistil length and ovule number were reduced. PIN1 expression was also lower in the triple mutant and the phenotypes could not be rescued by exogenous cytokinin application. pin1 complementation studies using genomic PIN1 constructs showed that the pistil phenotypes were only rescued when the PCRE1 domain, to which CRFs bind, was present. Without this domain, pin mutants resemble the crf2 crf3 crf6 triple mutant, indicating the pivotal role of CRFs in auxin-cytokinin crosstalk.

Angiosperms form the largest group of land plants and display an astonishing diversity of floral structures. The development of flowers greatly contributed to the evolutionary success of the angiosperms as they guarantee efficient reproduction with the help of either biotic or abiotic vectors. The female reproductive part of the flower is the gynoecium (also called pistil). Ovules arise from meristematic tissue within the gynoecium. Upon fertilization, these ovules develop into seeds while the gynoecium turns into a fruit. Gene regulatory networks involving transcription factors and hormonal communication regulate ovule primordium initiation, spacing on the placenta, and development. Ovule number and gynoecium size are usually correlated and several genetic factors that impact these traits have been identified. Understanding and fine-tuning the gene regulatory networks influencing ovule number and pistil length open up strategies for crop yield improvement, which is pivotal in light of a rapidly growing world population. In this review, we present an overview of the current knowledge of the genes and hormones involved in determining ovule number and gynoecium size. We propose a model for the gene regulatory network that guides the developmental processes that determine seed yield.

In higher plants, the female germline is formed from the megaspore mother cell (MMC), a single cell in the premeiotic ovule. Previously, it was reported that mutants in the RNA-dependent DNA methylation (RdDM) pathway might be involved in restricting the female germline to a single nucellus cell. We show that the DRM methyltransferase double mutant drm1drm2 also presents ectopic enlarged cells, consistent with supernumerary MMC-like cells. In wild-type ovules, MMC differentiation requires SPOROCYTELESS/NOZZLE (SPL/NZZ), as demonstrated by the spl/nzz mutant failing to develop an MMC. We address the poorly understood upstream regulation of SPL/NZZ in ovules, showing that the RdDM pathway is important to restrict SPL/NZZ expression. In ago9, rdr6 and drm1drm2 mutants, SPL/NZZ is expressed ectopically, suggesting that the multiple MMC-like cells observed might be attributable to the ectopic expression of SPL/NZZ. We show that the ovule identity gene, SEEDSTICK, directly regulates AGO9 and RDR6 expression in the ovule and therefore indirectly regulates SPL/NZZ expression. A model is presented describing the network required to restrict SPL/NZZ expression to specify a single MMC.