Genetic and Phenotypic Analysis of Ovule Development in Arabidopsis.

The plant seed is a remarkable structure that represents the single most important energy source in global diets. The stages of reproductive growth preceding seed formation are particularly important since they influence the number, size, and quality of seed produced. The progenitor of the seed is the ovule, a multicellular organ that produces a female gametophyte while maintaining a range of somatic ovule cells to protect the seed and ensure it receives maternal nourishment. Ovule development has been well characterized in Arabidopsis using a range of molecular, genetic, and cytological assays. These can provide insight into the mechanistic basis for ovule development, and opportunities to explore its evolutionary conservation. In this chapter, we describe some of these methods and tools that can be used to investigate early ovule development and cell differentiation.

The RNA-dependent DNA methylation pathway is required to restrict SPOROCYTELESS/NOZZLE expression to specify a single female germ cell precursor in Arabidopsis.

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.

Translating auxin responses into ovules, seeds and yield: Insight from Arabidopsis and the cereals.

Grain production in cereal crops depends on the stable formation of male and female gametes in the flower. In most angiosperms, the female gamete is produced from a germline located deep within the ovary, protected by several layers of maternal tissue, including the ovary wall, ovule integuments and nucellus. In the field, germline formation and floret fertility are major determinants of yield potential, contributing to traits such as seed number, weight and size. As such, stimuli affecting the timing and duration of reproductive phases, as well as the viability, size and number of cells within reproductive organs can significantly impact yield. One key stimulant is the phytohormone auxin, which influences growth and morphogenesis of female tissues during gynoecium development, gametophyte formation, and endosperm cellularization. In this review we consider the role of the auxin signaling pathway during ovule and seed development, first in the context of Arabidopsis and then in the cereals. We summarize the gene families involved and highlight distinct expression patterns that suggest a range of roles in reproductive cell specification and fate. This is discussed in terms of seed production and how targeted modification of different tissues might facilitate improvements.