What Makes a Grass? DROOPING LEAF Influences Flower and Leaf Development in Rice (2024)

The grass family, which includes >600 genera and 10,000 species, is one of the largest families of monocotyledonous plants and also one of the most ecologically and economically important of all plant families. Grasses and other monocots have a characteristic leaf and flower morphology that is distinct from dicotyledonous plants. Grass leaves exhibit a parallel leaf venation pattern, in contrast with the typical reticulate venation of dicot leaves, and there is a central vein, or midrib, that provides structural support to the leaf. Grass flowers are arranged in compound inflorescences called spikelets, and the individual florets do not have petals or sepals like most eudicot flowers. Instead, two bag-shaped lodicules are located at one side of the inner whorls containing carpels and stamens, and these are subtended by palea and lemma (bract-like structures that enclose the floral organs).

Regulation of floral organ specification is a central problem in plant development. The ABC model of floral organ specification (Bowman et al., 1991; Coen and Meyerowitz, 1991) accurately describes the basic mechanisms of floral pattern formation in eudicots, although it has been modified and expanded upon (Jack, 2001; Theissen, 2001; Lohman and Weigel, 2002). Much of the work to build and refine this model has been performed in Arabidopsis (Arabidopsis thaliana) and Antirrhinum majus. Work in monocot grasses such as maize (Zea mays) and rice (Oryza sativa) has shown that there is some conservation of floral homeotic gene function between monocots and dicots (reviewed in Goto et al., 2001). For example, maize silky1 and rice SUPERWOMAN1 encode apparent functionally equivalent orthologs of APETALA3 (AP3), a B-class gene that regulates specification of petals and stamens in Arabidopsis. However, our understanding of the molecular regulation of flower development in grasses and other monocots lags behind that in Arabidopsis and other eudicots.

In this issue of The Plant Cell, Yamaguchi et al. (pages 500–509) show that DROOPING LEAF (DL), which encodes a member of the plant-specific YABBY family of transcription factors, has an essential function in specifying carpel identity and meristem determinacy in rice flowers and also regulates midrib formation in rice leaves (Figure 1). Coen and Meyerowitz (1991) wrote that plant form is largely a function of meristem behavior. The results of Yamaguchi et al. show that the genetic mechanisms for carpel specification and meristem determinacy in monocots are distinct from that used by dicots and suggest that DL plays an important role in determining the characteristic form of monocots.

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Figure 1.

dl Mutant Flower Phenotype and Expression Pattern of DL in Wild-Type Rice.

Mutant dl-sup1 flower shows complete transformation of carpels into stamens (top left). Expression of DL in wild-type rice flower is restricted to the carpel primordia region (bottom left). Cross-section of mutant dl-sup1 leaf shows the lack of accumulation of cells along the adaxial-abaxial axis, corresponding to the lack of midrib formation (top right). Expression pattern of DL in wild-type rice leaf occurs in a band along the adaxial-abaxial axis and coincides with cell proliferation in the central region, which gives rise to the midrib structure (bottom right).

Nagasawa et al. (2003) previously described the phenotype of four recessive dl mutants. Yamaguchi et al. isolated five new dl alleles and cloned the gene using a combination of map-based cloning and sequencing of a mutant allele with a TOS17 retrotransposon insertion. The most severe dl alleles exhibited complete transformation of carpels into stamens, similar to the previously described superman (sup) mutant of Arabidopsis (Schultz et al., 1991; Bowman et al., 1992), and thus were called dl-sup. Floral organs appeared nearly normal in the weakest alleles, whereas intermediate alleles showed partial transformation of carpels and pleiotropic abnormalities, including production of multiple carpels and stamens and the formation of cell clusters. The similarity to sup was found to be largely superficial. SUP encodes a C2H2-type zinc finger protein, which appears to function mainly in establishing the boundary between the stamens and carpels by regulating cell proliferation in the third and fourth whorls (Sakai et al., 1995). Thus, mutations in SUP cause the formation of extra stamens in whorl 3 and inhibition of carpel formation in whorl 4 but do not cause homeotic transformation of carpels into stamens as in the dl mutants. In addition, all of the dl mutants exhibited defects in leaf midrib formation, resulting in the characteristic drooping leaf phenotype.

DL is a member of the plant-specific YABBY gene family, characterized by the presence of the YABBY region and a zinc finger–like domain (Bowman and Smyth, 1999). The YABBY region has partial similarity to the HMG domain, the defining feature of a family of DNA binding proteins called high mobility group (HMG) proteins, which are found in all eukaryotes. HMG proteins bind DNA non-sequence specifically and have the ability to bend and introduce supercoils into the double-stranded DNA helix. They are believed to work in concert with various sequence-specific DNA binding proteins and to facilitate the formation of nucleoprotein complexes important in processes such as transcription regulation, DNA repair, replication, and recombination (Grasser, 1998). The HMG domain consists of three α-helices that create an L-shaped fold that binds DNA nonspecifically but recognizes certain distorted DNA structures such as supercoiled DNA, four-way junctions, and DNA minicircles. Kanaya et al. (2002) found that the Arabidopsis YABBY protein FILAMENTOUS FLOWER (described in Sawa et al., 1999), binds to DNA in a non-sequence specific manner. However, the YABBY region is missing one of the three HMG α-helices, suggesting that YABBY proteins contact DNA in a somewhat different manner from HMG proteins. Bowman and Smyth (1999) suggested that the YABBY domain could represent part of an ancestral HMG gene that was combined with a zinc finger motif and co-opted for another function early in plant evolutionary history.

Most of the YABBY proteins that have been identified play roles in the development of floral organs. Many of these proteins are expressed in asymmetric patterns and are closely associated with asymmetric development of plant organs. For example, Arabidopsis INNER NO OUTER is expressed specifically on one side of ovule primordia in cells that give rise to the outer integument and is thought to be necessary for polarity determination in the central part of the ovule (Villanueva et al., 1999). FILAMENTOUS FLOWER is expressed only at the abaxial side of leaf and floral organ primordia and is necessary for normal development of both leaf and floral organs and for the maintenance of meristem activity (Sawa et al., 1999). DL is most similar to and a putative ortholog of Arabidopsis CRABS CLAW (CRC), which regulates carpel and nectary development in this species. Although both proteins are associated with carpel development, there are important differences in their apparent functions, which may relate to some of the fundamental differences in flower and leaf development between monocots and dicots.

In wild-type Arabidopsis flowers, CRC is expressed in carpels and in nectaries, the nectar-producing organs that arise at the base of the stamens during the latter stages of flower development. CRC plays a key role in nectary specification because crc mutant flowers show no sign of nectary development (however, constitutive expression of CRC does not induce ectopic nectaries in transgenic plants, suggesting that another factor is also required). Carpels develop in the normal position in crc mutant flowers, but the gynoecium shows developmental defects and an occasional extra carpel, suggesting the CRC functions to guide proper development of the gynoecium and may influence meristem determinacy but is not a primary factor in specifying carpel identity (Bowman and Smyth, 1999). The MADS box gene AGAMOUS is the primary C-class gene that specifies stamen and carpel identity in Arabidopsis (Bowman et al., 1991), although triple and quadruple mutant studies show that CRC and another gene, SPATULA, are partly involved in carpel specification (Alvarez and Smyth, 1999).

By contrast, loss-of-function mutations in DL caused complete homeotic transformation of carpels into stamens in rice flowers, indicating that it is necessary for the specification of carpel identity and can be considered a C-class gene in rice. In addition, DL was found to strongly influence meristem determinacy, as shown by the indeterminate formation of ectopic stamens in severe dl alleles, the production of multiple carpels and clusters of undifferentiated cells in intermediate dl alleles, and the expression pattern of a molecular marker of meristematic indeterminate cells in rice, OSH1. Finally, the characteristic drooping leaf phenotype of all dl alleles indicated that DL is required for midrib formation in rice leaves. Microscopic examination of midrib structure in wild-type and mutant plants suggested that DL regulates midrib formation by inducing cell proliferation in the leaf central region.

Most YABBY proteins are associated with abaxial/adaxial polarity specification, which is largely a function of their asymmetric expression patterns and, thus, ultimately dependent on the factors that regulate their expression. Bowman and Smyth (1999) showed that CRC is negatively regulated in whorls 1 and 2 by the A-class genes AP2 and LEUNIG and in whorl 3 by the B-class genes AP3 and PISTILLATA. Yamaguchi et al. show that DL similarly is negatively regulated by the AP3 ortholog SUPERWOMAN1. There appears to be limited interaction between CRC and the C-class gene AG in Arabidopsis (Bowman and Smyth, 1999). In rice, the AG ortholog OsMADS3 does not appear to be required for carpel identity specification, as shown by antisense suppression in transgenic plants (Kang et al., 1998). However, rice has at least one other AG ortholog whose function might overlap that of OsMADS3 (T. Yamaguchi and H.-Y. Hirano, unpublished data). It will be of considerable interest to determine the functions of the MADS-box C-class genes in rice and their interaction with DL.

Yamaguchi et al. concluded that there is fundamental conservation of function between CRC and DL, but DL acquired additional functions specifying carpel identity and midrib formation during evolution of the grasses. Thus, divergent evolution of the CRC/DL genes may underlie some of the major differences in leaf and flower form between monocots and dicots.

Notes

Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.ITI.

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