SMALL GRAIN 11 Controls Grain Size, Grain Number and Grain Yield in Rice

Abstract

Background: Grain size is one of key agronomic traits that determine grain yield in rice. Several regulators of grain size have been identified in rice, but the mechanisms that determine grain size and yield remain largely unknown.

Results: Here we characterize a small grain (smg11) mutant in rice, which exhibits small grains, dense panicles and the increased number of grains per panicle. Cloning and sequence analyses of the SMG11 gene reveal that smg11 is a new allele of DWARF2 (D2), which encodes a cytochrome P450 (CYP90D2) involved in brassinosteroid biosynthetic pathway. Overexpression of D2/SMG11 increases grain size and grain weight of wild-type plants. Overexpression of D2/SMG11 at a suitable level also significantly increases grain yield in rice. Cellular analyses indicate that D2/SMG11 controls grain size by promoting cell expansion. Further results reveal that D2/SMG11 influences expression of several known grain size genes involved in the regulation of cell expansion, revealing a novel link between D2/SMG11 and known grain size genes.

Conclusions: SMG11 controls grain size by promoting cell expansion in grain hulls. SMG11 regulates cell expansion, at least in part, by influencing expression of several grain size genes involved in the regulation of cell expansion. The smg11 is a new allele of DWARF2/D2. The suitable expression of SMG11 increases grain size, grain weight and grain yield. Our findings reveal the functions of D2/SMG11 in grain size and grain yield, suggesting that the suitable expression of D2/SMG11 is a promising approach to improve grain yield in rice.

Keywords: Cell expansion; Grain size; Rice; SMG11/D2.

Fig. 1

The smg11 mutant produces small grains. a Mature paddy rice grains of KYJ and smg11. b Brown rice grains of KYJ and smg11. c The average length of KYJ and smg11 grains. d The average width of KYJ and smg11 grains. e The 1000-grain weight of KYJ and smg11. Values c-e are given as mean ± SD. *P < 0.05; **P < 0.01 compared with the wild type by Student’s t-test. Bars: 2 mm in a and b

Fig. 2

smg11 produces dense and erect panicles. a KYJ (left) and smg11 (right) plants at the mature stage. b c Panicles of KYJ (left) and smg11 (right) at the mature stage. d Distance between panicle neck and each primary branch of KYJ and smg11 panicles. e The average length of KYJ and smg11 panicles. f Total number of KYJ and smg11 primary branches. g Total number of KYJ and smg11 secondary branches. h Grain number per panicle in KYJ and smg11. Values d-h are given as the mean ± SD. *P < 0.05; **P < 0.01 compared with the wild type by Student’s t-test. Bars: 20 cm in a; 5 cm in b-c

Fig. 3

smg11 influences cell expansion. a b SEM analysis of the outer surface of KYJ a and smg11 b lemmas. c The average length of outer epidermal cells in KYJ and smg11 lemmas. d The average width of outer epidermal cells in KYJ and smg11 lemmas. e f SEM analysis of the inner surface of KYJ e and smg11 f lemmas. g The average length of inner epidermal cells in KYJ and smg11 lemmas. h The average width of inner epidermal cells in KYJ and smg11 lemmas. i Expression levels of the indicated genes in KYJ and smg11 panicles. j Expression levels of the indicated genes in KYJ and smg11 panicles. Values c-d; g–j are means ± SD. *P < 0.05; **P < 0.01 compared with the wild type by Student’s t-test. Bars: 100 μm in a, b, e, f

Fig. 4

Identification of the D2/SMG11 gene. a The D2/SMG11 gene structure. The white boxes indicate the exons, and black lines indicate the introns. The start codon (ATG) and the stop codon (TAG) are indicated. The smg11 mutation in the SMG11/D2 gene is shown. b The dCAPS marker is developed according to the smg11 mutation. c The D2/SMG11 protein contains a membrane anchor domain (N1), a proline-rich domain (N2), a dioxygen binding domain (N3), a steroid binding domain (N4) and a heme binding domain (N5). d e Grains of KYJ, smg11, pActin: SMG11;smg11#1 and pActin: SMG11;smg11#2. pActin: SMG11;smg11 is smg11 transformed with pActin: SMG11. f g h Grain length f grain width g and 1000-grain weight h of KYJ, smg11, pActin: SMG11;smg11#1 and pActin: SMG11;smg11#2. i The number of KYJ, smg11, pActin: SMG11;smg11#1 and pActin: SMG11;smg11#2 secondary panicle branches. j Grain number per panicle of KYJ, smg11, pActin: SMG11;smg11#1 and pActin: SMG11;smg11#2. k Quantitative real-time RT-PCR analysis of SMG11/D2 gene expression in developing panicles of 1 cm (YP1) to 20 cm (YP20). Values f–k are given as mean ± SD. *P < 0.05; **P < 0.01 compared with smg11 by Student’s t-test. Bars: 2 mm in d-e

Fig. 5

Expression levels of BR-related genes in KYJ and smg11 panicles. Quantitative real-time RT-PCR analysis of BR-related gene expression in young panicles. Values are means ± SD. *P < 0.05 **P < 0.01 compared with the wild type by Student’s t-test

Fig. 6

Overexpression of D2/SMG11 increases grain size due to large cells in spikelet hulls. a b Grains of ZH11, pActin: SMG11#1, pActin: SMG11#2 and pActin: SMG11#3. pActin: SMG11 is ZH11 transformed with pActin: SMG11. c d SEM analysis of the outer surface of ZH11 c and pActin: SMG11#2 d lemmas. e, f SEM analysis of the inner surface of ZH11 e and pActin: SMG11#2 f lemmas. g h i Grain length g grain width h and 1000-grain weight i of ZH11, pActin:SMG11#1, pActin:SMG11#2 and pActin:SMG11#3. j Expression levels of SMG11/D2 in ZH11, pActin:SMG11#1, pActin:SMG11#2 and pActin:SMG11#3 panicles. k The average length of outer epidermal cells in ZH11 and pActin:SMG11#2 lemmas. l The average width of outer epidermal cells in ZH11 and pActin:SMG11#2 lemmas. m The average length of inner epidermal cells in ZH11 and pActin:SMG11#2 lemmas. n The average width of inner epidermal cells in ZH11 and pActin:SMG11#2 lemmas. Values g–n are given as mean ± SD. *P < 0.05 **P < 0.01 compared with ZH11 by Student’s t-test. Bars: 2 mm in a-b 100 μm in c–f

Fig. 7

The suitable expression of SMG11 increases grain yield. a Grain yield per plant of ZH11, pActin:SMG11#1, pActin:SMG11#2 and pActin:SMG11#3. b–d The primary panicle branch number b the secondary panicle branch number c and grain number per panicle d of ZH11, pActin:SMG11#1, pActin:SMG11#2 and pActin:SMG11#3. Values a–d are given as mean ± SD. *P < 0.05 **P < 0.01 compared with ZH11 by Student’s t test