Identification of a New OsBRI1 Weak Allele and Analysis of its Function in Grain Size Control

doi:10.11983/CBB19239

Abstract

Abstract:

Rice (Oryza sativa) grain size and grain weight are key agronomic traits that affect rice yield. Cloning and study of grain size genes are helpful to increase rice production. In order to further understand the mechanism of rice grain size control, a set of mutants with altered grain size from an EMS-treated elite japonica cultivar KYJ (Kuanyejing) were isolated. smg12 exhibits small grains, short plants, and reduced number of primary branches and secondary branches. Genetic analyses show that the smg12 mutant phenotypes are controlled by a single recessive gene. Our celluar analyses show that the small grain size phenotype of smg12 is caused by the decrease in cell size of glumes, indicating that SMG12 affects cell expansion. By using the Mutmap method, we reveal that the candidate gene for SMG12 is OsBRI1, which encodes a brassinolide receptor kinase. The smg12 mutant causes a substitution of the 2 074^th base (C to T) in OsBRI1, which results in an amino acid change (proline to serine). Therefore, this study identified a new mutant allele of OsBRI1 and provides a cellular and molecular basis for BR-mediated grain size control in rice.

Key words: rice, grain size, cell expansion, Mutmap, OsBRI1

Liurong Guan, Zupei Liu, Ran Xu, Penggen Duan, Guozheng Zhang, Haiyue Yu, Jing Li, Yuehua Luo, Yunhai Li. Identification of a New OsBRI1 Weak Allele and Analysis of its Function in Grain Size Control[J]. Chinese Bulletin of Botany, 2020, 55(3): 279-286.

Figures/Tables 6

Figure 1 Analysis of rice smg12 mutant phenotypes (A) KYJ (wild type) and smg12 mutant phenotype (Bar=10 cm); (B) Grains of KYJ and smg12 (Bars=1 cm); (C) Grain width; (D) Grain length; (E) Number of primary branches; (F) Number of secondary branches; (G) Plant height; (H) Thousand seed weight. Significance is determined using t-test, * indicates significant differences at P<0.05, ** indicates significant differences at P<0.01.

Table 1 Primer and enzyme for dCAPS analysis

Primer name	Primer sequence (5'→3')	DNA fragment length (bp)	Enzyme
smg12-1	CTTTCTCGGCACTTTCCTTG	154	HphI
smg12-1	CTATGGTCACATGGTGGCGGTG	154	HphI

Figure 2 SMG12 regulates cell size in grain hulls of rice (A) Outer epidermal cell number in the grain-length direction of the glume; (B) Outer epidermal cell number in the grain-width direction of the glume; (C) The longitudinal length of a single cell in the glume; (D) Transverse width of a single cell in the glume; (E) Cytological analysis diagram (Bars= 0.1 mm). ** indicates extremely significant difference between the mutant and the wild type (P<0.01).

Table 2 Genetic analysis of the rice mutant smg12

Hybrid combinations	Phenotype of F₁	F₂ generation			χ²_3:1
Hybrid combinations	Phenotype of F₁	Wild-phenotypic plant number	Mutant-phenotypic plant number	Total number	χ²_3:1
smg12/KYJ	Wild type	168	52	220	0.151

Table 3 Analysis of the candidate SNPs for the smg12 mutant

Chromosomal position	Physical location (bp)	Genotype (KYJ/smg12)	Gene location	Gene locus name	Frequency of sequencing (KYJ/smg12)	Mutation type
Chr. 1	15686971	G/A	Upstream of the gene	LOC_Os01g28040	0/6	/
Chr. 1	27750517	G/A	In the gene compartment	LOC_Os01g48420; LOC_Os01g48430	0/12	/
Chr. 1	28534819	G/A	Upstream and downstream of the gene	LOC_Os01g49630; LOC_Os01g49614	0/14	/
Chr. 1	29397508	G/A	In the gene compartment	LOC_Os01g51140; LOC_Os01g51154	0/12	/
Chr. 1	29788896	G/A	Upstream of the gene	LOC_Os01g51810	0/17	/
Chr. 1	29929259	G/A	Exon	LOC_Os01g52050	0/19	Changes in amino acids
Chr. 1	33892012	T/A	Upstream of the gene	LOC_Os01g58620	0/1	/
Chr. 1	39503252	G/A	Downstream of the gene	LOC_Os01g67980	0/11	/
Chr. 1	30392222	G/A	Upstream of the gene	LOC_Os01g52840; LOC_Os01g52851	0/5	/
Chr. 1	33745383	G/A	Intron	LOC_Os01g58400	0/8	/
Chr. 3	19627588	C/A	Upstream of the gene	LOC_Os03g35390	0/6	/
Chr. 6	17710551	G/A	Intron	LOC_Os06g30610	0/8	/

Figure 3 Identification of candidate genes (A) The dCAPS marker was developed to detect the smg12 mutation; (B) Population linkage analysis of the candidate gene LOC_Os01g52050; (C) The LOC_Os01g52050 gene structure, open boxes show the 5' and 3' untranslated regions, the closed box shows the coding sequence, and the start codon (ATG), the stop codon (TAG) and the LOC_Os01g52050 mutation site (C/T) are indicated; (D) Schematic of the LOC_Os01g52050 protein. P/S indicated the LOC_Os01g52050 mutation site.

References 30

[1]	宫李辉, 高振宇, 马伯军, 钱前 (2011). 水稻粒形遗传的研究进展. 植物学报 46, 597-605.
[2]	侯雷平, 李梅兰 (2001). 油菜素内酯(BR)促进植物生长机理研究进展. 植物学通报 18, 560-566.
[3]	Abe A, Kosugi S, Yoshida K, Natsume S, Takagi H, Kanzaki H, Matsumura H, Yoshida K, Mitsuoka C, Tamiru M, Innan H, Canno L, Kamoun S, Terauchi R (2012). Genome sequencing reveals agronomically important loci in rice using Mutmap. Nat Biotechnol 30, 174-178. URL PMID
[4]	Fan CC, Xing YZ, Mao HL, Lu TT, Han B, Xu CG, Li XH, Zhang QF (2006). GS3, a major QTL for grain length and weight and minor QTL for grain width and thickness in rice, encodes a putative transmembrane protein. Theor Appl Genet 112, 1164-1171. URL PMID
[5]	James MG, Denyer K, Myers AM (2003). Starch synthesis in the cereal endosperm. Curr Opin Plant Biol 6, 215-222.
[6]	Li N, Xu R, Duan PG, Li YH (2018). Control of grain size in rice. Plant Reprod 31, 237-251.
[7]	Li N, Xu R, Li YH (2019). Molecular networks of seed size control in plants. Annu Rev Plant Biol 70, 435-463. URL PMID
[8]	Liu LC, Tong HN, Xiao YH, Che RH, Xu F, Hu B, Liang CZ, Chu JF, Li JY, Chu CC (2015a). Activation of Big Grain1 significantly improves grain size by regulating auxin transport in rice. Proc Natl Acad Sci USA 112, 11102-11107. URL PMID
[9]	Liu SY, Hua L, Dong SJ, Chen HQ, Zhu XD, Jiang JE, Zhang F, Li YH, Fang XH, Chen F (2015b). OsMAPK6, a mitogen-activated protein kinase, influences rice grain size and biomass production. Plant J 84, 672-681. DOI URL PMID
[10]	Mao HL, Sun SY, Yao JL, Wang CR, Yu SB, Xu CG, Li XH, Zhang QF (2010). Linking differential domain functions of the GS3 protein to natural variation of grain size in rice. Proc Natl Acad Sci USA 107, 19579-19584.
[11]	Meng XZ, Zhang SQ (2013). MAPK cascades in plant disease resistance signaling. Annu Rev Phytopathol 51, 245-266. URL PMID
[12]	Miura K, Ashikari M, Matsuoka M (2011). The role of QTLs in the breeding of high-yielding rice. Trends Plant Sci 16, 319-326. URL PMID
[13]	Morinaka Y, Sakamoto T, Inukai Y, Agetsuma M, Kitano H, Ashikari M, Matsuoka M (2006). Morphological alteration caused by brassinosteroid insensitivity increases the biomass and grain production of rice. Plant Physiol 141, 924-931.
[14]	Nakamura A, Fujioka S, Sunohara H, Kamiya N, Hong Z, Inukai Y, Miura K, Takatsuto S, Yoshida S, Ueguchi- Tanaka M, Hasegawa Y, Kitano H, Matsuoka M (2006). The role of OsBRI1 and its homologous genes, OsBRL1 and OsBRL3, in rice. Plant Physiol 140, 580-590. DOI URL PMID
[15]	Sakamoto T, Matsuoka M (2008). Identifying and exploiting grain yield genes in rice. Curr Opin Plant Biol 11, 209-214. DOI URL PMID
[16]	Shomura A, Izawa T, Ebana K, Ebitani T, Kanegae H, Konishi S, Yano M (2008). Deletion in a gene associated with grain size increased yields during rice domestication. Nat Genet 40, 1023-1028. URL PMID
[17]	Si LZ, Chen JY, Huang XH, Gong H, Luo JH, Hou QQ, Zhou TY, Lu TT, Zhu JJ, Shangguan YY, Chen EW, Gong CX, Zhao Q, Jing YF, Zhao Y, Li Y, Cui LL, Fan DL, Lu YQ, Weng QJ, Wang YC, Zhan QL, Liu KY, Wei XH, An K, An G, Han B (2016). OsSPL13 controls grain size in cultivated rice. Nat Genet 48, 447-456.
[18]	Song XJ, Huang W, Shi M, Zhu MZ, Lin HX (2007). A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase. Nat Genet 39, 623-630. DOI URL PMID
[19]	Takagi H, Tamiru M, Abe A, Yoshida K, Uemura A, Yaegashi H, Obara T, Oikawa K, Utsushi H, Kanzaki E, Mitsuoka C, Natsume S, Kosugi S, Kanzaki H, Matsumura H, Urasaki N, Kamoun S, Terauchi R (2015). Mutmap accelerates breeding of a salt-tolerant rice cultivar. Nat Biotechnol 33, 445-449. URL PMID
[20]	Tanabe S, Ashikari M, Fujioka S, Takatsuto S, Yoshida S, Yano M, Yoshimura A, Kotano H, Matsuoka M, Fujisawa Y, Kato H, Lwasaki Y (2005). A novel cytochrome P450 is implicated in brassinosteroid biosynthesis via the characterization of a rice dwarf mutant, dwarf11, with reduced seed length. Plant Cell 17, 776-790. DOI URL PMID
[21]	Tanaka A, Nakagawa H, Tomita C, Shimatani Z, Ohtake M, Nomura T, Jiang CJ, Dubozet JG, Kikuchi S, Sekimoto H, Yokota T, Asami T, Kamakura T, Mori M (2009). BRASSINOSTEROID UPREGULATED1, encoding a helix-loop-helix protein, is a novel gene involved in brassinosteroid signaling and controls bending of the lamina joint in rice. Plant Physiol 151, 669-680. DOI URL PMID
[22]	Tong HN, Liu LC, Jin Y, Du L, Yin YH, Qian Q, Zhu LH, Chu CC (2012). DWARF AND LOW-TILLERING acts as a direct downstream target of a GSK3/SHAGGY-like kinase to mediate brassinosteroid responses in rice. Plant Cell 24, 2562-2577. URL PMID
[23]	Wang SK, Wu K, Yuan QB, Liu XY, Liu ZB, Lin XY, Zeng RZ, Zhu HT, Dong GJ, Qian Q, Zhang GQ, Fu XD (2012). Control of grain size, shape and quality by OsSPL16 in rice. Nat Genet 44, 950-954. DOI URL PMID
[24]	Xing YZ, Zhang QF (2010). Genetic and molecular bases of rice yield. Annu Rev Plant Biol 61, 421-442.
[25]	Xu R, Duan PG, Yu HY, Zhou ZK, Zhang BL, Wang RC, Li J, Zhang GZ, Zhuang SS, Lyu J, Li N, Chai TY, Tian ZX, Yao SG, Li YH (2018a). Control of grain size and weight by the OsMKKK10-OsMKK4-OsMAPK6 signaling pathway in rice. Mol Plant 11, 860-873.
[26]	Xu R, Yu HY, Wang JM, Duan PG, Zhang BL, Li J, Li Y, Xu JS, Lyu J, Li N, Chai TY, Li YH (2018b). A mitogen-activated protein kinase phosphatase influences grain size and weight in rice. Plant J 95, 937-946. DOI URL PMID
[27]	Zhang C, Bai MY, Chong K (2014). Brassinosteroid-mediated regulation of agronomic traits in rice. Plant Cell Rep 33, 683-696.
[28]	Zhao JF, Wu CX, Yuan SJ, Yin L, Sun W, Zhao QL, Zhao BH, Li XY (2013). Kinase activity of OsBRI1 is essential for brassinosteroids to regulate rice growth and development. Plant Sci 199-200, 113-120. DOI URL PMID
[29]	Zhou SR, Yin LL, Xue HW (2013). Functional genomics based understanding of rice endosperm development. Curr Opin Plant Biol 16, 236-246. URL PMID
[30]	Zuo JR, Li JY (2014). Molecular genetic dissection of quantitative trait loci regulating rice grain size. Annu Rev Genet 48, 99-118.