G1/ELE Functions in the Development of Rice Lemmas in Addition to Determining Identities of Empty Glumes

Abstract

Rice empty glumes, also named sterile lemmas or rudimentary lemmas according to different interpretations, are distinct from lemmas in morphology and cellular pattern. Consistently, the molecular mechanism to control the development of lemmas is different from that of empty glumes. Rice LEAFY HULL STERILE1(OsLHS1) and DROOPING LEAF(DL) regulate the cellular pattern and the number of vascular bundles of lemmas respectively, while LONG STERILE LEMMA1 (G1)/ELONGATED EMPTY GLUME (ELE) and PANICLE PHYTOMER2 (PAP2)/OsMADS34 determine identities of empty glumes. Though some progress has been made, identities of empty glumes remain unclear, and genetic interactions between lemma genes and glume genes have been rarely elucidated. In this research, a new G1/ELE mutant g1-6 was identified and the phenotype was analyzed. Similar to previously reported mutant lines of G1/ELE, empty glumes of g1-6 plants transform into lemma-like organs. Furthermore, Phenotypes of single and double mutant plants suggest that, in addition to their previously described gene-specific functions, G1/ELE and OsLHS1 play redundant roles in controlling vascular bundle number, cell volume, and cell layer number of empty glumes and lemmas. Meanwhile, expression patterns of G1/ELE in osmads1-z flowers and OsLHS1 in g1-6 flowers indicate they do not regulate each other at the level of transcription. Finally, down-regulation of the empty glume gene OsMADS34/PAP2 and ectopic expression of the lemma gene DL, in the g1-6 plants provide further evidence that empty glumes are sterile lemmas. Generally, our findings provided valuable information for better understanding functions of G1 and OsLHS1 in flower development and identities of empty glumes.

Keywords: G1/ELE; OsLHS1; empty glume; lemma; rice.

FIGURE 1

Positional cloning of g1–6 and sequence analysis. (A) The candidate gene was mapped in BAC AP003829 on the short arm of Chromosome 7. (B) Schematic representation of G1 and mutation site of g1–6. (C) Full cDNA of g1–6. The translation is terminated prematurely at the mutation site.

FIGURE 2

Phenotypes of spikelets and empty glumes. (A) One spikelet of wild type. (B) One g1–6 spikelet. (C) One empty glume of wild type. (D) One g1–6 empty glume. Transverse sections of one empty glume (E) and one lemma of wild type (F). Transverse sections of one g1–6 empty glume (G) and one g1–6 lemma (H). Red asterisks in (F–H) indicate vascular bundles. Bars = 2 mm in (A–D), 100 μm in (E), 200 μm in (F–H).

FIGURE 3

Scanning electron microscopy observation of empty glumes, lemmas, and paleas. Epidermis of one empty glume (A), one lemma (B), and one palea (C) of wild- type plants. Epidermis of one empty glume (D), one lemma (E), and one palea (F) of g1–6 plants. Bars = 50 μm.

FIGURE 4

Flower Phenotypes of osmads1-z and osmads1-z g1–6 plants. (A,B) Spikelets of osmads1-z. (C–E) Spikelets of osmads1-z g1–6; the lemma and palea were removed in (E) to show the inner organs. Epidermal cells of one empty glume (F), one lemma (G) and one palea (H) of osmads1-z; Epidermal cells of one leaf (I) and one lodicule (J) of wild type plants. Epidermal cells of one empty glume (K), one lemma (L) and one palea (M) of g1–6 osmads1-z. (N), Epidermal cells of one lodicule of osmads1-z. (O) Epidermal cells of one lodicule of osmads1-z g1–6. Bars = 2 mm in (A–E), 50 μm in (F–O) except for (I), 100 μm in (I).

FIGURE 5

Statistical analyses of number of vascular bundles in empty glumes, lemmas, and paleas. For every kind of organs, 15 samples of wild-type (WT), g1–6, osmads1-z, g1–6 osmads1-z were examined with transverse sections, respectively. The means were statistically analyzed with the Duncan test and grouped (A, B, and C) according to significant differences at P < 0.01. Error bars indicate SD.

FIGURE 6

Histological observation of empty glumes, lemmas, and paleas. Transverse sections of one empty glume (A), one lemma (B), and one palea (C) in wild type plants. Transverse sections of one abnormal empty glume (D), one lemma (E), and one palea (F) in g1–6 plants. Transverse sections of one empty glume (G), one lemma (H), and one palea (I) in osmads1-z plants; Transverse sections of one empty glume (J), one lemma (K), and one palea (L) in osmads1-z g1–6 plants. Sc, silicified cells; fs, fibrous sclerenchyma; spc, spongy parenchymatous cells; nsc, no silicified cells. Bars = 50 μm.

FIGURE 7

Statistical analyses of the thickness of empty glumes, lemmas, and paleas. For every kind of organ, 13 samples of wild type, 14 samples of g1–6, 17 samples of osmads1-z, 22 samples of g1–6 osmads1-z plants were measured with transverse sections, respectively. The means were statistically analyzed with the Duncan test and grouped (A, B, and C) according to significant differences at P < 0.01. Error bars indicate SD.

FIGURE 8

Expression patterns of DL, OsLHS1, OsMADS34/PAP2, and G1/ELE. (A) Expression analyses of DL gene in empty glumes, lemmas, and paleas. (B) Expression analyses of OsLHS1 in empty glumes, lemmas, and paleas. (C) Expression analyses of G1/ELE in empty glumes, lemmas, and paleas. (D) Expression analyses of OsMADS34/PAP2 in empty glumes, lemmas, and paleas. Error bars indicate SD.