SEMI-ROLLED LEAF1 Encodes a Putative Glycosylphosphatidylinositol-Anchored Protein and Modulates Rice Leaf Rolling by Regulating the Formation of Bulliform Cells

Jing-Jing Xiang,Guang-Heng Zhang,Hong-Wei Xue,Qian Qian

doi:10.1104/pp.112.199968

Abstract

Leaf rolling is an important agronomic trait in rice (Oryza sativa) breeding and moderate leaf rolling maintains the erectness of leaves and minimizes shadowing between leaves, leading to improved photosynthetic efficiency and grain yields. Although a few rolled-leaf mutants have been identified and some genes controlling leaf rolling have been isolated, the molecular mechanisms of leaf rolling still need to be elucidated. Here we report the isolation and characterization of SEMI-ROLLED LEAF1 (SRL1), a gene involved in the regulation of leaf rolling. Mutants srl1-1 (point mutation) and srl1-2 (transferred DNA insertion) exhibit adaxially rolled leaves due to the increased numbers of bulliform cells at the adaxial cell layers, which could be rescued by complementary expression of SRL1. SRL1 is expressed in various tissues and is expressed at low levels in bulliform cells. SRL1 protein is located at the plasma membrane and predicted to be a putative glycosylphosphatidylinositol-anchored protein. Moreover, analysis of the gene expression profile of cells that will become epidermal cells in wild type but probably bulliform cells in srl1-1 by laser-captured microdissection revealed that the expression of genes encoding vacuolar H(+)-ATPase (subunits A, B, C, and D) and H(+)-pyrophosphatase, which are increased during the formation of bulliform cells, were up-regulated in srl1-1. These results provide the transcript profile of rice leaf cells that will become bulliform cells and demonstrate that SRL1 regulates leaf rolling through inhibiting the formation of bulliform cells by negatively regulating the expression of genes encoding vacuolar H(+)-ATPase subunits and H(+)-pyrophosphatase, which will help to understand the mechanism regulating leaf rolling.

Highlights

National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 200032 Shanghai, China (J.-J.X., H.-W.X.); and State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006, Zhejiang, China (G.-H.Z., Q.Q.)
SEMI-ROLLED LEAF1 (SRL1) Negatively Regulates the Formation of Bulliform Cells and Modulates Leaf Rolling. In grass species such as rice, leaf rolling is induced by water loss from bulliform cells on the leaf upper epidermis (O’toole and Cruz, 1980), the number and density of bulliform cells may affect the extent of leaf rolling
Srl1-1 and srl1-2 mutants displayed adaxially rolled leaves resulted from increased number of bulliform cells on the adaxial side of leaf blades, which may substantiate the role of bulliform cells in controlling leaf rolling

Summary

RESULTS

Srl, a Rice Mutant with Adaxially Rolled Leaves, Has the Increased Bulliform Cells at the Adaxial Epidermis. Observation revealed the increased bulliform cells at the adaxial epidermis of the third leaf blade in srl compared with that of the wild type at seedling stage (Fig. 1C; Supplemental Fig. S1). Phenotypic observation and cross section of leaf blades of the transgenic plants showed the flat leaves and similar number of bulliform cells as wild-type plants (Fig. 2, C and D), demonstrating that complementary expression of SRL1 rescued the leafrolling phenotype of srl and the crucial role of SRL1 in leaf-rolling control. Observation of GFP fluorescence by transiently expressing SRL1-GFP fusion protein (GFP was inserted at 364 amino acids upstream of the v-site as the N-terminal signal peptide and C-terminal propeptide of SRL1 might be cut off for GPI processing) in the protoplasts of Arabidopsis and rice, and onion (Allium cepa) epidermal cells, respectively, revealed that SRL1-GFP localizes predominantly at the plasma membrane (Fig. 4C; Supplemental Fig. S5). Further qRT-PCR analysis confirmed the increased expression levels of genes encoding vacuolar H+-pyrophosphatase and H+-ATPase (subunits A, B, C, and D) in srl (Fig. 5C), suggesting that the formation of bulliform cells may be coupled to increased expression of genes encoding vacuolar H+ATPase subunits and H+-pyrophosphatase

DISCUSSION

MATERIALS AND METHODS