SeeDev-binary@ldeleger:SeeDev-binary-11573014-1 JSONTXT

The Arabidopsis LEAFY COTYLEDON2 (LEC2) gene is a central embryonic regulator that serves critical roles both early and late during embryo development. LEC2 is required for the maintenance of suspensor morphology, specification of cotyledon identity, progression through the maturation phase, and suppression of premature germination. We cloned the LEC2 gene on the basis of its chromosomal position and showed that the predicted polypeptide contains a B3 domain, a DNA-binding motif unique to plants that is characteristic of several transcription factors. We showed that LEC2 RNA accumulates primarily during seed development, consistent with our finding that LEC2 shares greatest similarity with the B3 domain transcription factors that act primarily in developing seeds, VIVIPAROUS1/ABA INSENSITIVE3 and FUSCA3. Ectopic, postembryonic expression of LEC2 in transgenic plants induces the formation of somatic embryos and other organ-like structures and often confers embryonic characteristics to seedlings. Together, these results suggest that LEC2 is a transcriptional regulator that establishes a cellular environment sufficient to initiate embryo development. Embryogenesis in flowering plants begins with the double fertilization event in which the zygote and endosperm are formed after fusion of sperm cells with the egg cell and central cell of the female gametophyte, respectively. The endosperm initially undergoes syncytial development with formation of nuclear-cytoplasmic domains, but later cellularizes (1). The developing embryo is nourished by the endosperm and, in many plants, only the peripheral layer of the endosperm remains in the mature seed. Development of the zygote into the mature embryo can be divided conceptually into two distinct phases. During the early morphogenesis phase, the basic body plan of the plant is established with expression of polarity as a shoot–root axis, specification of morphological domains within the embryo, and formation of embryonic tissue and organ systems (2–4). The morphogenesis phase is followed by a period of maturation in which processes critical for seed formation occur (5, 6). During this late phase, reserves such as storage proteins and lipids are synthesized at high rates and accumulate in the seed. It is also during the maturation phase that the embryo acquires the ability to withstand desiccation at the final stage of seed development. At the end of embryogenesis, the seed consists of a mature, desiccated embryo that is quiescent metabolically. Although many aspects of embryogenesis have been characterized extensively, little is known at a mechanistic level of the processes that initiate embryo development. The Arabidopsis LEAFY COTYLEDON (LEC) genes, LEC1, LEC2, and FUSCA3 (FUS3), play key roles in controlling embryo development (7). Unlike most other embryonic regulators that function during specific stages of embryogenesis (8–11), LEC genes are unique in that they are required for normal development during both the morphogenesis and maturation phases. Early in embryogenesis, LEC genes are required to specify suspensor cell fate and cotyledon identity (12–16). Late in embryogenesis, LEC genes are needed during the maturation phase for the acquisition of desiccation tolerance and the expression of many maturation-specific genes (13–17). Consistent with the finding that conditions that promote maturation suppress germination (6), lec mutant embryos prematurely activate the postgermination program (13, 15, 16, 18). Thus, LEC genes play a central role in controlling many aspects of embryogenesis, and they are candidates as regulators that coordinate the morphogenesis and maturation phases. Identification and analysis of two LEC genes confirmed their regulatory role in embryogenesis and provided insight into their functions. LEC1 shares extensive sequence similarity with the HAP3 subunit of CCAAT-binding transcription factor, implicating LEC1 as a transcriptional regulator (12). Ectopic expression of LEC1 confers embryonic characteristics to seedlings and results in the formation of embryo-like structures on the surfaces of leaves, suggesting that the gene plays a role in conferring embryogenic competence to cells (12). Thus, we hypothesized that LEC1 establishes a cellular environment that promotes embryo development and that this environment coordinates the morphogenesis and maturation phases. FUS3 also encodes a regulatory protein: a B3 domain transcription factor that accumulates primarily during seed development (19). Transient assays showed that FUS3 is sufficient to activate genes usually expressed during maturation (20). Thus, two LEC genes seem to be involved in controlling embryo development by regulating transcription of other genes. In this article, we focus on the LEC2 gene to determine its role in embryo development. Because genetic studies suggest that LEC1 and LEC2 may have partially redundant functions (12, 15), it is possible that LEC2 also functions in the initiation and coordination of embryo development. We cloned the LEC2 gene and showed that it is expressed preferentially during embryogenesis and encodes a protein with similarity to other seed-specific transcription factors. Significant insight into the role of the gene was obtained by showing that transgenic plants expressing the LEC2 gene ectopically form somatic embryos. Together, these results indicate that LEC2 is sufficient to induce embryogenic competence.

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