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Rice appearance quality
Rice appearance quality
11
Hao Zhou, Peng Yun, Yuqing He National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, China
In rice, traits related to appearance quality have a large impact on market value and play a pivotal role in the adoption of new varieties. However, different grain quality traits are prized by different local cultures and cuisines, unlike other cereals such as wheat, barley, and maize that are sold largely in processed forms, the physical properties of rice grains are immediately obvious to consumers (Fitzgerald et al., 2009). On the other hand, the appearance quality of rice is closely linked to rice yield, and when breeders are breeding new varieties, they tend to combine high quality with yield, unlike focusing only on yield, which they used to do (Zuo and Li, 2014). Thus, genetic investigation of rice appearance quality is of vital importance for not only improving the market value but also the breeding efficiency of rice.
1. Definition, classification, and diversity of rice appearance quality A rice grain is composed of hull (lemma and palea), bran layers (pericarp, aleurone, subaleurone layers), embryo, and endosperm. The most common bran layer for rice is brown, so it is called “brown rice” when removing the hull (Fig. 11.1A). The outer layer of natural brown bran is stripped off to create “white rice,” removing most of the fiber, vitamins, minerals, and amino acids. When bran layers and embryo are removed during the polishing process, it will become “white rice” and also the general consumed form of rice (Fig. 11.1B). The physical properties of rice include grain size and shape, chalkiness, translucence, and color. Grain size includes grain length, width, and thickness, and grain shape is determined by grain length and width. In most cases, indica rice has a long and slender shape and japonica rice has a short and round shape, and in many cases, short or/and slender grains are preferred, such as the short slender variety Samba Mahsuri, which is extremely popular in Southern India (Sundaram et al., 2008). The edible part of rice is the endosperm and is mainly composed of starch, and chalky grain results from the opaque spots in the endosperm (Fig. 11.1C). Chalkiness can be categorized as white core, white belly, and white back on the basis of the location of opaque spots in endosperm. Chalkiness rate directly affects consumer
Rice. https://doi.org/10.1016/B978-0-12-811508-4.00011-3 Copyright © 2019 AACCI. Published by Elsevier Inc. in cooperation with AACC International. All rights reserved.
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acceptance of rice, and as for nonwaxy rice, only those with a chalkiness rate lower than 5% will be defined as high-quality rice. The rice appearance quality is mainly determined by grain shape and chalkiness rate. The other two aspects, translucence and color, are influenced by composition of starch, chalkiness, and content of pigment (anthocyanin, flavonoids, and carotenoids) in rice. Cultivated rice (Oryza sativa) has a very wide genetic diversity, and the appearance quality of rice is the most direct characteristic of rice genetic diversity (Fig. 11.1). Rice, which was mainly produced in Asia and was planted worldwide, was influenced by different local environments, climate, and artificial selection, and has a large diversity of appearance. As shown in Fig. 11.1B, the grain size of rice has a dramatic diversity. In cultivated rice, the grain length ranges from 3 to 11 mm and width from 1.2 to 3.8 mm (Fitzgerald et al., 2009; Huang et al., 2013). Brown rice without hull can have many colors, and the most common varieties are red rice, black rice, brown rice, and wild rice (Sweeney et al., 2006; Furukawa et al., 2007; Oikawa et al., 2015). Since these brown rices have some special nutrition parts, they have their own market value. Polished rice mainly displays a white and transparent physical appearance, and varieties with high chalkiness or low amylose content may show a nontransparent phenotype. Chalkiness can be categorized as white core, white belly, and white back on the basis of the location of opaque spots in endosperm (Fig. 11.1C). Waxy rice has a white and opaque phenotype due to the extremely low amylose content (0%e2%). With the development of technology, scientists have bred polished rice with yellow and black colors through, respectively, synthesis of carotenoids and anthocyanin via transgenic methods (Ye et al., 2000; Paine et al., 2005; Zhu et al., 2017).
(A)
(B)
(C)
White belly
White core
White back
Figure 11.1 Genetic diversity of appearance quality. (A) Brown rice with different colors. (B) White rice with different grain shape and size. Scale bar represents 3 mm. (C) Three kinds of chalkiness in rice.
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2. Grain shape and rice yield Grain size related traits, including grain length, width, thickness, and length-to-width ratio, have a direct impact on grain weight and quality, further affecting the commercialization of rice (Fitzgerald et al., 2009; Huang et al., 2013; Zuo and Li, 2014). It is well known that rice yield is determined by three major components: number of panicles per plant, number of grains per panicle, and grain weight. Among these, grain weight is the most reliable trait, which is measured as 1000-grain weight. Thus, increasing grain weight by enlarging grain size will be an effective way to improve yield. On the other hand, grain size not only influences appearance but also affects the milling quality of rice. Rice grain will lose some parts during processing, and the longer or wider the grain is, the losses increase. Rice grain size has always been an important agronomic trait that is artificially selected during breeding for its influence on rice yield and quality (Shomura et al., 2008; Takano-Kai et al., 2009). Breeders tend to choose plants with large grain size to improve yield, while making sure that plants with such grain size have good milling quality and will meet the markets’ demands.
3. Effects of grain chalkiness on rice milling and eating quality Rice chalkiness is not only a major component of appearance but also a key factor that determines the milling quality and cooking and eating quality (Lanning et al., 2011; Bowles, 2012; Siebenmorgen et al., 2013). The starch granules in the chalky region of rice grain are not as tightly packed into cells as starch granules in the normal region, and are small and disorganized, resulting in the reduction of grain tenacity and the head rice rate. Thus, in the aspect of milling, chalkiness is an unfavorable trait. On the other hand, the higher the chalkiness rate, the worse the cooking and eating quality will be (Singh et al., 2003; Cheng et al., 2005). Since the chalky region has a higher transfer temperature and a lower gel consistency with a high hardness, the gelatinization temperature of chalky rice is much higher than normal rice. What’s more, researchers have observed that the increase of chalkiness rate will lead to a decrease of taste. Thus, chalkiness is an unfavorable trait both for production and for market demands. Reducing the chalkiness has become the common goal of high-quality rice breeding.
4. Genetic bases of rice grain shape Regarding the importance of grain appearance on rice commercialization, scientists have carried out numerous genetic studies on grain shape and chalkiness traits (Bao, 2014). The grain shape of rice is controlled by polygenes and has a relative high heredity, and is not easily influenced by environment. Over the past decades, hundreds of grain shape quantitative trait loci (QTLs) had been identified, and more than a dozen of them were cloned (Table 11.1), which laid the foundation of the elucidation of the
Table 11.1 A summary of major quantitative trait loci (QTLs) controlling grain shape and chalkiness Encoded protein
Nature of the beneficial alleles
Reference
GS3
Grain length
G-protein g subunit
165 bp C/A, premature stop
Fan et al. (2006), Takano-Kai et al. (2009), and Mao et al. (2010)
GW2
Grain width
RING-type E3 ubiquitin ligase
1 bp deletion, premature stop
Song et al. (2007)
GW5
Grain width
Calmodulin-binding protein
1212 bp InDel or 950 bp InDel, expression altered
Shomura et al. (2008), Weng et al. (2008), Duan et al. (2017), and Liu et al. (2017)
GS5
Grain width
Serine carboxypeptidase
1109 and 1032, expression altered
Li et al. (2011)
GL3
Grain length
Protein phosphatase
1092 bp C/A and 1495 bp C/T, expression altered
Qi et al. (2012)
GW8/SPL16
Grain width
Transcription factor
10 bp deletion, expression altered
Wang et al. (2012)
TGW6
Grain weight
IAA-glucose hydrolase
1 bp deletion, premature stop
Ishimaru et al. (2013)
GS2
Grain length
Growth-regulating factor
487 bp TC/AA, perturbing OsmiR396-directed regulation
Hu et al. (2015)
GL7
Grain shape
LONGIGOLIA protein
17.1 kb tandem duplication, expression
Wang et al. (2015)
GW7
Grain shape
TRM protein
11 bp deletion and 18 bp insertion, expression
Wang et al. (2015)
GLW7/SPL13
Grain shape
Plant-specific transcription factor
CATTC copies, expression altered
Si et al. (2016)
GIF1
Grain shape, grain chalkiness
Cell wall invertase
Loss of function
Wang et al. (2008)
Chalk5
Grain chalkiness
Vacuolar Hþ-translocating pyrophosphatase
expression altered, 721 and 485
Li et al. (2014)
Rice
Trait
374
Gene
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genetic basis of grain shape and provided valuable information for the improvement of rice appearance quality (Zuo and Li, 2014).
4.1
The importance of GS3 and GW5 in rice quality and yield
The sequencing of natural variation varieties and genome-wide association studies both confirmed that GS3 and GW5 are two major genes controlling rice grain length and grain width, respectively (Takano-Kai et al., 2009; Gong et al., 2017; Liu et al., 2017). The first cloned rice grain length gene, GS3, is located on the third chromosome and encodes a G-protein g subunit (Fan et al., 2006). GS3 negatively regulates grain length, and the C to A transition in the second exon leads to premature transcription termination as well as the improvement of grain length. GW5, which is located on the fifth chromosome of rice, encodes a calmodulin-binding protein and negatively regulates rice grain width (Liu et al., 2017). In round grain varieties, the upstream deletion (1212bp in indica and 950bp in japonica) reduces the expression level of GW5 and increases the grain width of rice. Both the mutant genotypes (gw5 and gs3) of GW5 and GS3 improve the grain weight of rice, but gw5 greatly promotes the chalkiness rate of rice (Huang et al., 2015; Gong et al., 2017). Thus, the functional GW5 and nonfunctional gs3 formed a genotype combination, GW5gs3, balanced the quality and yield of rice, and was selected in rice breeding process. And the GW5gs3 was also reported to be a beneficial genotype combination for the outcrossing rate of male sterile lines and assumed to be important in hybrid rice breeding (Zhou et al., 2017).
4.2
Other QTLs in rice grain shape studies
Besides GS3 and GW5, there are many other QTLs that contribute to the natural variation of rice grain shape (Table 11.1). GW2 is a major QTL for grain width and weight, and a rare mutation of GW2 results in increased cell number and acceleration of grain filling (Song et al., 2007). The GW2 was first cloned by using a BC3F2 population derived from WY3 and Fengaizhan. GW2 encodes a previously unknown RINGtype E3 ubiquitin ligase, and a rare 1-bp deletion in the GW2 gene of WY3 results in a premature stop codon in its exon 4, causing the wide-grain phenotype in WY3. Sequencing of natural varieties found that the rare nonfunctional variation of GW2 mainly exists in japonica. GS5 is the first cloned positive regulator of grain size, which encodes a putative serine carboxypeptidase and is a minor QTL for grain width (Li et al., 2011). Polymorphisms in the promoter of GS5 may lead to different expression levels of GS5, and a higher expression of GS5 may increase grain width and weight due to increased cell proliferation and expansion in spikelet hulls. Researchers later found that GS5 can competitively inhibit the interaction between OsBAK1-7 and OsMSBP1, which suggests that GS5 might control grain size by influencing BR signaling (Xu et al., 2015). GW8 is another major QTL for grain width and weight and encodes an SQUAMOSA promoter-binding protein-like 16 (OsSPL16), which belongs to the protein family of SBP domain-containing transcription factors (Wang et al., 2012). A 10-bp deletion in the promoter region of GW8 has been shown to be responsible for the grain size variation and grain width increases with the increasing
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expression level of GW8. GL7/GW7 positively controls grain length and negatively controls grain width (Wang et al., 2015a,b). It has been reported that GL7/GW7 can not only increase grain size but also improve rice quality without influencing tie yield of rice (Wang et al., 2015a,b). A newly identified grain length gene, GL3.3,encoding a GSK3/SHAGGY-like kinase, epistatically interacts with GS3 to form extra-long grains in rice (Xia et al., 2017). Numerous genetic research studies of grain shape shed light on uncovering the genetic basis of grain shape. Combining beneficial grain shape genes such as gs3, GW5, gw8, and GL/GW7 will offer a good opportunity for greatly improving grain shape.
4.3
Molecular mechanisms regulating grain shape
The analyses of grain shape QTLs have provided important clues about the molecular mechanisms that regulate this key agronomic trait. Currently available evidence suggests that grain size is controlled by three major signal pathway phytohormones, including ubiquitination-mediated proteasomal degradation, and G-protein signaling pathways (Fig. 11.2). The phytohormones pathway is the major pathway for molecular mechanism of grain shape. Phytohormones like auxin, cytokinin, and brassinosteroid (BR) all participate in the regulation of grain shape. TGW6, a major QTL that controls rice grain weight and grain filling, encodes an IAA-glucosehydrolase and plays an
D61
Brassinosteroid ? ? GS5 D11
?
D1/RGA1 RGB1
?
SMG1
SRS5 GS2
GW5 BU1
G-protein sigaling
? qGL3
GLW7 ?
GSK2 ?
TGW6 BG1
OsCKX2
Auxin
DST LP
Cytokinin
GS6
Grain shape
GW7 GL7
GW8
? GS3 DEP1
Proteasomal degradation IPA1 GW2
Figure 11.2 A schematic representation illustrating major regulatory genes that control seed size. These genes are involved in the regulation of cell division and cell expansion, presumably via different signaling pathways (indicated by shaded areas) or via unknown pathways (GS5, GW8, and IPA1). For conciseness, only components that are known to be involved in the control of seed size are shown in each signaling pathway. This figure was updated based on Zuo and Li’s studies (Zuo and Li, 2014).
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important role in the regulation of auxin homeostasis during endosperm development (Ishimaru et al., 2013). The regulatory mechanism of BR on seed development has been vigorously investigated in recent years. The major grain width gene, GW5, which encodes a calmodulin-binding protein, was found to interact with GSK2 (glycogen synthase kinase 2), which is a negative regulator of BR signaling and a homologue of Arabidopsis BIN2 (brassinosteroid insensitive 2), and to mediate BR-responsive gene expression and growth responses (Liu et al., 2017). GS6, GS2, and GL3.1 are also genes regulating grain shape through the BR signal pathway. GW2 encodes a RING-type E3 ubiquitin ligase and is found to regulate grain width through proteasomal degradation pathway mediated by ubiquitin (Song et al., 2007). GS3 is reported to be functioning in G-protein signaling, and loss of function of this putative g subunit of G-protein increases grain size (Takano-Kai et al., 2009; Mao et al., 2010). These achievements made on the molecular mechanism of grain shape will be beneficial for the improvement of grain shape and will make the manipulation of grain shape possible.
5. Genetic bases of rice grain chalkiness Unlike grain shape, chalkiness is a typical multigene-controlled trait, which is sensitive to the environment and regulated through complex genetic pathways. It has been reported that the high temperature during grain-filling stage will induce the formation of chalkiness, and even an extreme floury phenotype, which remains a giant obstacle for investigating chalkiness (Ishimaru et al., 2009; Lanning et al., 2011). Until now, only a few chalkiness QTLs have been fine mapped for the low heredity of chalkiness, and only one gene has been cloned (Table 11.1), leading to the genetic basis and molecular mechanism of chalkiness remaining unknown.
5.1
QTL mapping for chalkiness in rice
Numerous QTLs for chalkiness or related components have been mapped hitherto by different populations, including doubled-haploid (DH) population, F2, recombinant inbred line (RIL) population, and chromosome segment substitution line population. Peng et al. (2014) resolved the genetic basis of chalkiness components using five populations (two DH lines and three RILs) across two environments. A total of 79 QTLs were detected, and 36.1% of them were consistently detected in the two environments. These indicated that chalkiness is genetically controlled and affected largely by environment. Zhao et al. (2016) detected 78 and 43 QTLs for grain chalkiness in nine environments by two sets of RILs from reciprocal crosses between Lemont and Teqing. Among the QTLs, 14 and 5 were stably expressed across different environments. Yun et al. (2016) mapped 5 QTLs for both white back rate (WBR) and white core rate using a RIL population. Two QTLs for WBR (qWBR2 and qWBR5) showed similar chromosomal locations with GW2 and qSW5/GW5, which have been cloned previously to control the grain width and chalkiness (Huang et al., 2015; Gong et al., 2017).
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5.2
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The cloning of Chalk5 and identification of other QTLs
By contrast, only a few QTLs responsible for grain chalkiness have been finely mapped and functionally characterized. Chalk5, the only cloned and functionally characterized chalkiness gene, was located on chromosome 5 and encodes a vacuolar Hþtranslocating pyrophosphatase. Li et al. (2014) found that Chalk5 expressed specially in endosperm is a positive regulator of grain white belly rate, and the ZS97 allele greatly increased chalkiness rate in near-isogenic lines and transgenic lines (Fig. 11.3B). Two SNPs in the promoter region of chalk5 repressed the expression level of Chalk5, resulting in a decrease of chalkiness. Sequencing of cultivated rice and genome-wide association analysis further revealed that these two SNPs are the main reason for the variation of chalkiness in indica. Besides chalk5, there are many starch synthesis genes and grain shape genes related to grain chalkiness. Loss of function of Flo5/SSIIIa, which participates in the elongation of short-chain starch, will influence the structure of starch, and T-DNA insertion of Flo5 displays a white core phenotype. It has been thought that there is a significant positive relation between grain width and chalkiness, and many cloned grain width genes do indeed influence grain chalkiness.
5.3
Molecular mechanisms of chalkiness formation
Due to the lack of cloned chalkiness-related genes and research of the molecular mechanism, the cause of chalkiness is poorly understood. As understood from existing research results, chalkiness is formed by the loose arrangement between starch granules and proteasome in endosperm during grain-filling stage and might be a result of different accumulation rates of nutrition parts due to the different transport channels of photosynthesis products. Ishimaru et al. (2009) tried to figure out the cause of chalkiness through heat stress and found that high temperature can induce the formation of chalk. They further proved that the abnormal loss of water of endosperm under high temperature was the main cause of white core and floury phenotype. The research of Li et al. (2014) demonstrated that elevated expression of Chalk5 increases chalkiness rate by disturbing the pH homeostasis in the endomembrane trafficking system in developing seeds, which affects the biogenesis of protein bodies and increases the small vesicle-like structures, leading to the formation of air spaces among endosperm storage substances and resulting in chalky grains (Fig. 11.3A and C).
6.
Genetic improvement of rice appearance quality
Development of new cultivars with improved appearance quality is important for rice breeding. The identification and cloning of appearance quality QTLs provide gene sources for marker-assisted selection (MAS) and genetic engineering. Fan et al. (2006) cloned the major grain length gene GS3 and developed a functional molecular marker, SF28, according to the C to A variation. Wang et al. (2012) utilized SF28 for the MAS of GS3, to pyramid the nonfunctional allele gs3 and gw8 into HJX74, a
Rice appearance quality
(A)
379
(B)
Golgi apparatus Nucleus
Prevacuolar compartment
Zhenshan 97
OHH+ OHH+ H + H2 O H2 O
Chalk5 Water loss
H94
(C)
Vacuole (proteosome) NIL(H94)
NIL(ZS97)
OX(–)
OX(+)
ZpZc(–)
ZpZc(+)
Figure 11.3 Chalk5 increase grain chalkiness in rice. (A) A proposed working model for Chalk5. Chalk5 accelerates the water loss of vacuole and induces the accumulation of small vesicle-like structures, leading to the formation of air spaces among endosperm storage substances, resulting in chalky grains. (B) Grain chalkiness of zhenshan 97, H94, NIL(ZS97), NIL(H94), and the two transformants in T2 progeny. OX(þ) and ZpZc(þ) indicate grains from T2 plants expressing the coding region of Chalk5 from Zhenshan 97 driven by the 35S promoter and the native promoter from Zhenshan 97, respectively. OX() and ZpZc() represent the corresponding transgene-negative segregants. (C) Scanning electron microscopy images of transverse sections from the endosperm bellies of mature seeds. Positions marked by rectangles in the middle bellies of the endosperm were analyzed by scanning electron microscopy. Scale bars: 500 mm (top), 10 mm (middle), 5 mm (bottom).
variety with short and wide grains. They successfully improved the grain shape of HJX74 and formed a similar grain shape to Basimati385, a variety with slender grain. MAS of superior appearance quality genes was also applied to hybrid rice breeding. QTL pyramiding of the GW7 and gs3 alleles has helped the development of new high-yielding indica hybrid rice varieties (for example, Taifengyou 55 and Taifengyou 208) with simultaneously improved yield and grain quality (Wang et al., 2015a,b). Unlike grain shape, little progress has been made in selecting against chalk over the years. Zhao et al. (2016) found that low chalk lines in the RIL population maintained their chalk classification across the nine environments. And introgression of five QTLs together almost eliminated grain chalkiness. CRISPR/Cas9 systems have been successfully used as efficient tools for genome editing in a variety of species. We can knock out negative regulators of grain quality efficiently to speed up the breeding process in rice. Li et al. (2016) used a CRISPR/ Cas9 system to edit four yield-related genes (Gn1a, DEP1, GS3, and IPA1) in Zhonghua 11 and obtained phenotypes similar to those of previously reported mutants, respectively. Another study targeted eight genes in rice (including GS3 and GW2) using a CRISPR/Cas9 multiplex genome editing system, and various phenotypes related to the editing genes were observed (Shen et al., 2017).
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Major issues and prospects in studies on rice appearance quality
Over the past decade, scientists have made impressive achievements in genomic studies on rice appearance quality. Those achievements can be summed up in three aspects: (1) the discovery of important genes for rice appearance quality; (2) the progress in molecular properties, expression and regulation of genes; (3) the application of MAS and transgenic technology. However, rice appearance quality is composed of many traits, and there were pervasive interactions among quality traits, between quality and yield, as well as between quality and environment. Additionally, the key scientific problem of rice appearance quality is not obvious to people. Those issues limit the improvement of rice appearance quality. Nonchalkiness is the primary goal in genetic improvement and basic research of rice appearance quality. Rice chalkiness is a trait conferring all the milling, appearance, eating, and cooking qualities. As the white rices of most varieties have approximately the same nutrition quality, the chalkiness tends to be the crucial problem need to be solved in rice quality improvement. The general high chalkiness rate of indica hybrid rice forms a common view about the negative correlation between rice quality and yield. However, few indica hybrid rice varieties are of both good quality and yield, which indicates that quality can coexist with yield if we can figure out their genetic mechanisms. Currently, the following measures are useful in reducing rice chalkiness: (1) properly improve grain length to width and break the adverse linkage between chalkiness QTLs and grain width QTLs; (2) take advantage of diverse natural germplasm resources and artificial mutant resources to identify and clone more chalkiness functional genes, and to ascertain the molecular, celled, physiological, and biochemical mechanism during chalkiness formation; (3) look for varieties with low chalkiness in heat stress and utilize their superior genes to solve the problem of chalkiness formation in high temperature. Sustained high temperatures after rice heading will greatly increase the proportion of chalky rice. In Asian rice growing areas, this climatic condition now occurs more often than it did in previous years, possibly due to global warming. Global warming will become a global issue in future agriculture. Thus, thorough and comprehensive genetic studies on rice quality are needed to face the challenges of future climate change.
References Bowles, D., 2012. Towards increased crop productivity and quality. Current Opinion in Biotechnology 23, 202e203. Cheng, F., Zhong, L., Wang, F., Zhang, G., 2005. Differences in cooking and eating properties between chalky and translucent parts in rice grains. Food Chemistry 90, 39e46. Duan, P., Xu, J., Zeng, D., Zhang, B., Geng, M., Zhang, G., Huang, K., Huang, L., Xu, R., Ge, S., 2017. Natural variation in the promoter of GSE5 contributes to grain size diversity in rice. Molecular Plant 10, 685e694.
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Fan, C., Xing, Y., Mao, H., Lu, T., Han, B., Xu, C., Li, X., Zhang, Q., 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. Theoretical and Applied Genetics 112, 1164e1171. Fitzgerald, M.A., McCouch, S.R., Hall, R.D., 2009. Not just a grain of rice: the quest for quality. Trends in Plant Science 14, 133e139. Furukawa, T., Maekawa, M., Oki, T., Suda, I., Iida, S., Shimada, H., Takamure, I., Kadowaki, K., 2007. The Rc and Rd genes are involved in proanthocyanidin synthesis in rice pericarp. The Plant Journal 49, 91e102. Gong, J., Jiashun, M., Yan, Z., Qiang, Z., Qi, F., Qilin, Z., Benyi, C., Junhui, X., Xuehui, H., Han, B., 2017. Dissecting the genetic basis of grain shape and chalkiness traits in hybrid rice using multiple collaborative populations. Molecular Plant 10, 1353e1356. Hu, J., Wang, Y., Fang, Y., Zeng, L., Xu, J., Yu, H., Shi, Z., Pan, J., Zhang, D., Kang, S., Zhu, L., Dong, G., Guo, L., Zeng, D., Zhang, G., Xie, L., Xiong, G., Li, J., Qian, Q., 2015. A rare allele of GS2 enhances grain size and grain yield in rice. Molecular Plant 8, 1455e1465. Huang, R., Jiang, L., Zheng, J., Wang, T., Wang, H., Huang, Y., Hong, Z., 2013. Genetic bases of rice grain shape: so many genes, so little known. Trends in Plant Science 18, 218e226. Huang, X., Yang, S., Gong, J., Zhao, Y., Feng, Q., Gong, H., Li, W., Zhan, Q., Cheng, B., Xia, J., 2015. Genomic analysis of hybrid rice varieties reveals numerous superior alleles that contribute to heterosis. Nature Communications 6, 6258. Ishimaru, T., Horigane, A.K., Ida, M., Iwasawa, N., San-oh, Y.A., Nakazono, M., Nishizawa, N.K., Masumura, T., Kondo, M., Yoshida, M., 2009. Formation of grain chalkiness and changes in water distribution in developing rice caryopses grown under high-temperature stress. Journal of Cereal Science 50, 166e174. Ishimaru, K., Hirotsu, N., Madoka, Y., Murakami, N., Hara, N., Onodera, H., Kashiwagi, T., Ujiie, K., Shimizu, B-i, Onishi, A., 2013. Loss of function of the IAA-glucose hydrolase gene TGW6 enhances rice grain weight and increases yield. Nature Genetics 45, 707e711. Lanning, S.B., Siebenmorgen, T.J., Counce, P.A., Ambardekar, A.A., Mauromoustakos, A., 2011. Extreme nighttime air temperatures in 2010 impact rice chalkiness and milling quality. Field Crops Research 124, 132e136. Li, Y., Fan, C., Xing, Y., Jiang, Y., Luo, L., Sun, L., Shao, D., Xu, C., Li, X., Xiao, J., He, Y., Zhang, Q., 2011. Natural variation in GS5 plays an important role in regulating grain size and yield in rice. Nature Genetics 43, 1266e1269. Li, Y., Fan, C., Xing, Y., Yun, P., Luo, L., Yan, B., Peng, B., Xie, W., Wang, G., Li, X., Xiao, J., Xu, C., He, Y., 2014. Chalk5 encodes a vacuolar H(þ)-translocating pyrophosphatase influencing grain chalkiness in rice. Nature Genetics 46, 398e404. Li, M., Li, X., Zhou, Z., Wu, P., Fang, M., Pan, X., Lin, Q., Luo, W., Wu, G., Li, H., 2016. Reassessment of the four yield-related genes Gn1a, DEP1, GS3, and IPA1 in rice using a CRISPR/Cas9 system. Frontiers in plant science 7, 377. Liu, J., Chen, J., Zheng, X., Wu, F., Lin, Q., Heng, Y., Tian, P., Cheng, Z., Yu, X., Zhou, K., 2017. GW5 acts in the brassinosteroid signalling pathway to regulate grain width and weight in rice. Nature Plants 3, 17043. Mao, H., Sun, S., Yao, J., Wang, C., Yu, S., Xu, C., Li, X., Zhang, Q., 2010. Linking differential domain functions of the GS3 protein to natural variation of grain size in rice. Proceedings of the National Academy of Sciences of the United States of America 107, 19579e19584. Oikawa, T., Maeda, H., Oguchi, T., Yamaguchi, T., Tanabe, N., Ebana, K., Yano, M., Ebitani, T., Izawa, T., 2015. The birth of a black rice gene and its local spread by introgression. The Plant Cell 27, 2401e2414.
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Paine, J.A., Shipton, C.A., Chaggar, S., Howells, R.M., Kennedy, M.J., Vernon, G., Wright, S.Y., Hinchliffe, E., Adams, J.L., Silverstone, A.L., 2005. Improving the nutritional value of Golden Rice through increased pro-vitamin A content. Nature Biotechnology 23, 482e487. Peng, B., Wang, L., Fan, C., Jiang, G., Luo, L., Li, Y., He, Y., 2014. Comparative mapping of chalkiness components in rice using five populations across two environments. BMC Genetics 15, 49. Qi, P., Lin, Y.S., Song, X.J., Shen, J.B., Huang, W., Shan, J.X., Zhu, M.Z., Jiang, L., Gao, J.P., Lin, H.X., 2012. The novel quantitative trait locus GL3.1 controls rice grain size and yield by regulating Cyclin-T1;3. Cell Research 22, 1666e1680. Shen, L., Hua, Y., Fu, Y., Li, J., Liu, Q., Jiao, X., Xin, G., Wang, J., Wang, X., Yan, C., 2017. Rapid generation of genetic diversity by multiplex CRISPR/Cas9 genome editing in rice. Science China Life Sciences 1e10. 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. Nature Genetics 40, 1023e1028. Si, L., Chen, J., Huang, X., Gong, H., Luo, J., Hou, Q., Zhou, T., Lu, T., Zhu, J., Shangguan, Y., 2016. OsSPL13 controls grain size in cultivated rice. Nature Genetics 48, 447e456. Siebenmorgen, T.J., Grigg, B.C., Lanning, S.B., 2013. Impacts of preharvest factors during kernel development on rice quality and functionality. Annual Review of Food Science and Technology 4, 101e115. Singh, N., Sodhi, N., Kaur, M., Saxena, S., 2003. Physico-chemical, morphological, thermal, cooking and textural properties of chalky and translucent rice kernels. Food Chemistry 82, 433e439. Song, X.-J., Huang, W., Shi, M., Zhu, M.-Z., Lin, H.-X., 2007. A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase. Nature Genetics 39, 623e630. Sundaram, R.M., Vishnupriya, M.R., Biradar, S.K., Laha, G.S., Reddy, G.A., Rani, N.S., Sarma, N.P., Sonti, R.V., 2008. Marker assisted introgression of bacterial blight resistance in Samba Mahsuri, an elite indica rice variety. Euphytica 160, 411e422. Sweeney, M.T., Thomson, M.J., Pfeil, B.E., McCouch, S., 2006. Caught red-handed: Rc encodes a basic helix-loop-helix protein conditioning red pericarp in rice. The Plant Cell 18, 283e294. Takano-Kai, N., Jiang, H., Kubo, T., Sweeney, M., Matsumoto, T., Kanamori, H., Padhukasahasram, B., Bustamante, C., Yoshimura, A., Doi, K., McCouch, S., 2009. Evolutionary history of GS3, a gene conferring grain length in rice. Genetics 182, 1323e1334. Wang, E., Wang, J., Zhu, X., Hao, W., Wang, L., Li, Q., Zhang, L., He, W., Lu, B., Lin, H., 2008. Control of rice grain-filling and yield by a gene with a potential signature of domestication. Nature Genetics 40, 1370e1374. Wang, S., Wu, K., Yuan, Q., Liu, X., Liu, Z., Lin, X., Zeng, R., Zhu, H., Dong, G., Qian, Q., Zhang, G., Fu, X., 2012. Control of grain size, shape and quality by OsSPL16 in rice. Nature Genetics 44, 950e954. Wang, S., Li, S., Liu, Q., Wu, K., Zhang, J., Wang, S., Wang, Y., Chen, X., Zhang, Y., Gao, C., 2015a. The OsSPL16-GW7 regulatory module determines grain shape and simultaneously improves rice yield and grain quality. Nature Genetics 47, 949e954. Wang, Y., Xiong, G., Hu, J., Jiang, L., Yu, H., Xu, J., Fang, Y., Zeng, L., Xu, E., Xu, J., Ye, W., Meng, X., Liu, R., Chen, H., Jing, Y., Wang, Y., Zhu, X., Li, J., Qian, Q., 2015b. Copy number variation at the GL7 locus contributes to grain size diversity in rice. Nature Genetics 47, 944e948.
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Weng, J., Gu, S., Wan, X., Gao, H., Guo, T., Su, N., Lei, C., Zhang, X., Cheng, Z., Guo, X., Wang, J., Jiang, L., Zhai, H., Wan, J., 2008. Isolation and initial characterization of GW5, a major QTL associated with rice grain width and weight. Cell Research 18, 1199e1209. Xia, D., Zhou, H., Liu, R., Dan, W., Li, P., Wu, B., Chen, J., Wang, L., Gao, G., Zhang, Q., He, Y., 2018. GL3. 3, a Novel QTL Encoding a GSK3/SHAGGY-like Kinase, Epistatically Interacts with GS3 to Produce Extra-long Grains in Rice. Molecular plant 11, 754e756. Xu, C., Liu, Y., Li, Y., Xu, X., Xu, C., Li, X., Xiao, J., Zhang, Q., 2015. Differential expression of GS5 regulates grain size in rice. Journal of Experimental Botany 66, 2611e2623. Yan, W.G., Bao, J.S., 2014. Genes and QTLs for rice grain quality improvement. RiceGermplasm. Genetics and Improvement (InTech), pp. 239e278. Ye, X., Al-Babili, S., Kl€oti, A., Zhang, J., Lucca, P., Beyer, P., Potrykus, I., 2000. Engineering the provitamin A (b-carotene) biosynthetic pathway into (carotenoid-free) rice endosperm. Science 287, 303e305. Yun, P., Zhu, Y., Wu, B., Gao, G., Sun, P., Zhang Q,He, Y., 2016. Genetic mapping and confirmation of quantitative trait loci for grain chalkiness in rice. Molecular Breeding 36, 162. Zhao, X., Daygon, V.D., McNally, K.L., Hamilton, R.S., Xie, F., Reinke, R.F., Fitzgerald, M.A., 2016. Identification of stable QTLs causing chalk in rice grains in nine environments. Theoretical and Applied Genetics 129, 141e153. Zhou, H., Li, P., Xie, W., Hussain, S., Li, Y., Xia, D., Zhao, H., Sun, S., Chen, J., Ye, H., 2017. Genome-wide association analyses reveal the genetic basis of stigma exsertion in rice. Molecular Plant 10, 634e644. Zhu, Q., Yu, S., Zeng, D., Liu, H., Wang, H., Yang, Z., Xie, X., Shen, R., Tan, J., Li, H., 2017. Development of “purple endosperm rice” by engineering anthocyanin biosynthesis in the endosperm with a high-efficiency transgene stacking system. Molecular Plant 10, 918e929. Zuo, J., Li, J., 2014. Molecular genetic dissection of quantitative trait loci regulating rice grain size. Annual Review of Genetics 48, 99e118.
11
Hao Zhou, Peng Yun, Yuqing He National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, China
In rice, traits related to appearance quality have a large impact on market value and play a pivotal role in the adoption of new varieties. However, different grain quality traits are prized by different local cultures and cuisines, unlike other cereals such as wheat, barley, and maize that are sold largely in processed forms, the physical properties of rice grains are immediately obvious to consumers (Fitzgerald et al., 2009). On the other hand, the appearance quality of rice is closely linked to rice yield, and when breeders are breeding new varieties, they tend to combine high quality with yield, unlike focusing only on yield, which they used to do (Zuo and Li, 2014). Thus, genetic investigation of rice appearance quality is of vital importance for not only improving the market value but also the breeding efficiency of rice.
1. Definition, classification, and diversity of rice appearance quality A rice grain is composed of hull (lemma and palea), bran layers (pericarp, aleurone, subaleurone layers), embryo, and endosperm. The most common bran layer for rice is brown, so it is called “brown rice” when removing the hull (Fig. 11.1A). The outer layer of natural brown bran is stripped off to create “white rice,” removing most of the fiber, vitamins, minerals, and amino acids. When bran layers and embryo are removed during the polishing process, it will become “white rice” and also the general consumed form of rice (Fig. 11.1B). The physical properties of rice include grain size and shape, chalkiness, translucence, and color. Grain size includes grain length, width, and thickness, and grain shape is determined by grain length and width. In most cases, indica rice has a long and slender shape and japonica rice has a short and round shape, and in many cases, short or/and slender grains are preferred, such as the short slender variety Samba Mahsuri, which is extremely popular in Southern India (Sundaram et al., 2008). The edible part of rice is the endosperm and is mainly composed of starch, and chalky grain results from the opaque spots in the endosperm (Fig. 11.1C). Chalkiness can be categorized as white core, white belly, and white back on the basis of the location of opaque spots in endosperm. Chalkiness rate directly affects consumer
Rice. https://doi.org/10.1016/B978-0-12-811508-4.00011-3 Copyright © 2019 AACCI. Published by Elsevier Inc. in cooperation with AACC International. All rights reserved.
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acceptance of rice, and as for nonwaxy rice, only those with a chalkiness rate lower than 5% will be defined as high-quality rice. The rice appearance quality is mainly determined by grain shape and chalkiness rate. The other two aspects, translucence and color, are influenced by composition of starch, chalkiness, and content of pigment (anthocyanin, flavonoids, and carotenoids) in rice. Cultivated rice (Oryza sativa) has a very wide genetic diversity, and the appearance quality of rice is the most direct characteristic of rice genetic diversity (Fig. 11.1). Rice, which was mainly produced in Asia and was planted worldwide, was influenced by different local environments, climate, and artificial selection, and has a large diversity of appearance. As shown in Fig. 11.1B, the grain size of rice has a dramatic diversity. In cultivated rice, the grain length ranges from 3 to 11 mm and width from 1.2 to 3.8 mm (Fitzgerald et al., 2009; Huang et al., 2013). Brown rice without hull can have many colors, and the most common varieties are red rice, black rice, brown rice, and wild rice (Sweeney et al., 2006; Furukawa et al., 2007; Oikawa et al., 2015). Since these brown rices have some special nutrition parts, they have their own market value. Polished rice mainly displays a white and transparent physical appearance, and varieties with high chalkiness or low amylose content may show a nontransparent phenotype. Chalkiness can be categorized as white core, white belly, and white back on the basis of the location of opaque spots in endosperm (Fig. 11.1C). Waxy rice has a white and opaque phenotype due to the extremely low amylose content (0%e2%). With the development of technology, scientists have bred polished rice with yellow and black colors through, respectively, synthesis of carotenoids and anthocyanin via transgenic methods (Ye et al., 2000; Paine et al., 2005; Zhu et al., 2017).
(A)
(B)
(C)
White belly
White core
White back
Figure 11.1 Genetic diversity of appearance quality. (A) Brown rice with different colors. (B) White rice with different grain shape and size. Scale bar represents 3 mm. (C) Three kinds of chalkiness in rice.
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2. Grain shape and rice yield Grain size related traits, including grain length, width, thickness, and length-to-width ratio, have a direct impact on grain weight and quality, further affecting the commercialization of rice (Fitzgerald et al., 2009; Huang et al., 2013; Zuo and Li, 2014). It is well known that rice yield is determined by three major components: number of panicles per plant, number of grains per panicle, and grain weight. Among these, grain weight is the most reliable trait, which is measured as 1000-grain weight. Thus, increasing grain weight by enlarging grain size will be an effective way to improve yield. On the other hand, grain size not only influences appearance but also affects the milling quality of rice. Rice grain will lose some parts during processing, and the longer or wider the grain is, the losses increase. Rice grain size has always been an important agronomic trait that is artificially selected during breeding for its influence on rice yield and quality (Shomura et al., 2008; Takano-Kai et al., 2009). Breeders tend to choose plants with large grain size to improve yield, while making sure that plants with such grain size have good milling quality and will meet the markets’ demands.
3. Effects of grain chalkiness on rice milling and eating quality Rice chalkiness is not only a major component of appearance but also a key factor that determines the milling quality and cooking and eating quality (Lanning et al., 2011; Bowles, 2012; Siebenmorgen et al., 2013). The starch granules in the chalky region of rice grain are not as tightly packed into cells as starch granules in the normal region, and are small and disorganized, resulting in the reduction of grain tenacity and the head rice rate. Thus, in the aspect of milling, chalkiness is an unfavorable trait. On the other hand, the higher the chalkiness rate, the worse the cooking and eating quality will be (Singh et al., 2003; Cheng et al., 2005). Since the chalky region has a higher transfer temperature and a lower gel consistency with a high hardness, the gelatinization temperature of chalky rice is much higher than normal rice. What’s more, researchers have observed that the increase of chalkiness rate will lead to a decrease of taste. Thus, chalkiness is an unfavorable trait both for production and for market demands. Reducing the chalkiness has become the common goal of high-quality rice breeding.
4. Genetic bases of rice grain shape Regarding the importance of grain appearance on rice commercialization, scientists have carried out numerous genetic studies on grain shape and chalkiness traits (Bao, 2014). The grain shape of rice is controlled by polygenes and has a relative high heredity, and is not easily influenced by environment. Over the past decades, hundreds of grain shape quantitative trait loci (QTLs) had been identified, and more than a dozen of them were cloned (Table 11.1), which laid the foundation of the elucidation of the
Table 11.1 A summary of major quantitative trait loci (QTLs) controlling grain shape and chalkiness Encoded protein
Nature of the beneficial alleles
Reference
GS3
Grain length
G-protein g subunit
165 bp C/A, premature stop
Fan et al. (2006), Takano-Kai et al. (2009), and Mao et al. (2010)
GW2
Grain width
RING-type E3 ubiquitin ligase
1 bp deletion, premature stop
Song et al. (2007)
GW5
Grain width
Calmodulin-binding protein
1212 bp InDel or 950 bp InDel, expression altered
Shomura et al. (2008), Weng et al. (2008), Duan et al. (2017), and Liu et al. (2017)
GS5
Grain width
Serine carboxypeptidase
1109 and 1032, expression altered
Li et al. (2011)
GL3
Grain length
Protein phosphatase
1092 bp C/A and 1495 bp C/T, expression altered
Qi et al. (2012)
GW8/SPL16
Grain width
Transcription factor
10 bp deletion, expression altered
Wang et al. (2012)
TGW6
Grain weight
IAA-glucose hydrolase
1 bp deletion, premature stop
Ishimaru et al. (2013)
GS2
Grain length
Growth-regulating factor
487 bp TC/AA, perturbing OsmiR396-directed regulation
Hu et al. (2015)
GL7
Grain shape
LONGIGOLIA protein
17.1 kb tandem duplication, expression
Wang et al. (2015)
GW7
Grain shape
TRM protein
11 bp deletion and 18 bp insertion, expression
Wang et al. (2015)
GLW7/SPL13
Grain shape
Plant-specific transcription factor
CATTC copies, expression altered
Si et al. (2016)
GIF1
Grain shape, grain chalkiness
Cell wall invertase
Loss of function
Wang et al. (2008)
Chalk5
Grain chalkiness
Vacuolar Hþ-translocating pyrophosphatase
expression altered, 721 and 485
Li et al. (2014)
Rice
Trait
374
Gene
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genetic basis of grain shape and provided valuable information for the improvement of rice appearance quality (Zuo and Li, 2014).
4.1
The importance of GS3 and GW5 in rice quality and yield
The sequencing of natural variation varieties and genome-wide association studies both confirmed that GS3 and GW5 are two major genes controlling rice grain length and grain width, respectively (Takano-Kai et al., 2009; Gong et al., 2017; Liu et al., 2017). The first cloned rice grain length gene, GS3, is located on the third chromosome and encodes a G-protein g subunit (Fan et al., 2006). GS3 negatively regulates grain length, and the C to A transition in the second exon leads to premature transcription termination as well as the improvement of grain length. GW5, which is located on the fifth chromosome of rice, encodes a calmodulin-binding protein and negatively regulates rice grain width (Liu et al., 2017). In round grain varieties, the upstream deletion (1212bp in indica and 950bp in japonica) reduces the expression level of GW5 and increases the grain width of rice. Both the mutant genotypes (gw5 and gs3) of GW5 and GS3 improve the grain weight of rice, but gw5 greatly promotes the chalkiness rate of rice (Huang et al., 2015; Gong et al., 2017). Thus, the functional GW5 and nonfunctional gs3 formed a genotype combination, GW5gs3, balanced the quality and yield of rice, and was selected in rice breeding process. And the GW5gs3 was also reported to be a beneficial genotype combination for the outcrossing rate of male sterile lines and assumed to be important in hybrid rice breeding (Zhou et al., 2017).
4.2
Other QTLs in rice grain shape studies
Besides GS3 and GW5, there are many other QTLs that contribute to the natural variation of rice grain shape (Table 11.1). GW2 is a major QTL for grain width and weight, and a rare mutation of GW2 results in increased cell number and acceleration of grain filling (Song et al., 2007). The GW2 was first cloned by using a BC3F2 population derived from WY3 and Fengaizhan. GW2 encodes a previously unknown RINGtype E3 ubiquitin ligase, and a rare 1-bp deletion in the GW2 gene of WY3 results in a premature stop codon in its exon 4, causing the wide-grain phenotype in WY3. Sequencing of natural varieties found that the rare nonfunctional variation of GW2 mainly exists in japonica. GS5 is the first cloned positive regulator of grain size, which encodes a putative serine carboxypeptidase and is a minor QTL for grain width (Li et al., 2011). Polymorphisms in the promoter of GS5 may lead to different expression levels of GS5, and a higher expression of GS5 may increase grain width and weight due to increased cell proliferation and expansion in spikelet hulls. Researchers later found that GS5 can competitively inhibit the interaction between OsBAK1-7 and OsMSBP1, which suggests that GS5 might control grain size by influencing BR signaling (Xu et al., 2015). GW8 is another major QTL for grain width and weight and encodes an SQUAMOSA promoter-binding protein-like 16 (OsSPL16), which belongs to the protein family of SBP domain-containing transcription factors (Wang et al., 2012). A 10-bp deletion in the promoter region of GW8 has been shown to be responsible for the grain size variation and grain width increases with the increasing
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expression level of GW8. GL7/GW7 positively controls grain length and negatively controls grain width (Wang et al., 2015a,b). It has been reported that GL7/GW7 can not only increase grain size but also improve rice quality without influencing tie yield of rice (Wang et al., 2015a,b). A newly identified grain length gene, GL3.3,encoding a GSK3/SHAGGY-like kinase, epistatically interacts with GS3 to form extra-long grains in rice (Xia et al., 2017). Numerous genetic research studies of grain shape shed light on uncovering the genetic basis of grain shape. Combining beneficial grain shape genes such as gs3, GW5, gw8, and GL/GW7 will offer a good opportunity for greatly improving grain shape.
4.3
Molecular mechanisms regulating grain shape
The analyses of grain shape QTLs have provided important clues about the molecular mechanisms that regulate this key agronomic trait. Currently available evidence suggests that grain size is controlled by three major signal pathway phytohormones, including ubiquitination-mediated proteasomal degradation, and G-protein signaling pathways (Fig. 11.2). The phytohormones pathway is the major pathway for molecular mechanism of grain shape. Phytohormones like auxin, cytokinin, and brassinosteroid (BR) all participate in the regulation of grain shape. TGW6, a major QTL that controls rice grain weight and grain filling, encodes an IAA-glucosehydrolase and plays an
D61
Brassinosteroid ? ? GS5 D11
?
D1/RGA1 RGB1
?
SMG1
SRS5 GS2
GW5 BU1
G-protein sigaling
? qGL3
GLW7 ?
GSK2 ?
TGW6 BG1
OsCKX2
Auxin
DST LP
Cytokinin
GS6
Grain shape
GW7 GL7
GW8
? GS3 DEP1
Proteasomal degradation IPA1 GW2
Figure 11.2 A schematic representation illustrating major regulatory genes that control seed size. These genes are involved in the regulation of cell division and cell expansion, presumably via different signaling pathways (indicated by shaded areas) or via unknown pathways (GS5, GW8, and IPA1). For conciseness, only components that are known to be involved in the control of seed size are shown in each signaling pathway. This figure was updated based on Zuo and Li’s studies (Zuo and Li, 2014).
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important role in the regulation of auxin homeostasis during endosperm development (Ishimaru et al., 2013). The regulatory mechanism of BR on seed development has been vigorously investigated in recent years. The major grain width gene, GW5, which encodes a calmodulin-binding protein, was found to interact with GSK2 (glycogen synthase kinase 2), which is a negative regulator of BR signaling and a homologue of Arabidopsis BIN2 (brassinosteroid insensitive 2), and to mediate BR-responsive gene expression and growth responses (Liu et al., 2017). GS6, GS2, and GL3.1 are also genes regulating grain shape through the BR signal pathway. GW2 encodes a RING-type E3 ubiquitin ligase and is found to regulate grain width through proteasomal degradation pathway mediated by ubiquitin (Song et al., 2007). GS3 is reported to be functioning in G-protein signaling, and loss of function of this putative g subunit of G-protein increases grain size (Takano-Kai et al., 2009; Mao et al., 2010). These achievements made on the molecular mechanism of grain shape will be beneficial for the improvement of grain shape and will make the manipulation of grain shape possible.
5. Genetic bases of rice grain chalkiness Unlike grain shape, chalkiness is a typical multigene-controlled trait, which is sensitive to the environment and regulated through complex genetic pathways. It has been reported that the high temperature during grain-filling stage will induce the formation of chalkiness, and even an extreme floury phenotype, which remains a giant obstacle for investigating chalkiness (Ishimaru et al., 2009; Lanning et al., 2011). Until now, only a few chalkiness QTLs have been fine mapped for the low heredity of chalkiness, and only one gene has been cloned (Table 11.1), leading to the genetic basis and molecular mechanism of chalkiness remaining unknown.
5.1
QTL mapping for chalkiness in rice
Numerous QTLs for chalkiness or related components have been mapped hitherto by different populations, including doubled-haploid (DH) population, F2, recombinant inbred line (RIL) population, and chromosome segment substitution line population. Peng et al. (2014) resolved the genetic basis of chalkiness components using five populations (two DH lines and three RILs) across two environments. A total of 79 QTLs were detected, and 36.1% of them were consistently detected in the two environments. These indicated that chalkiness is genetically controlled and affected largely by environment. Zhao et al. (2016) detected 78 and 43 QTLs for grain chalkiness in nine environments by two sets of RILs from reciprocal crosses between Lemont and Teqing. Among the QTLs, 14 and 5 were stably expressed across different environments. Yun et al. (2016) mapped 5 QTLs for both white back rate (WBR) and white core rate using a RIL population. Two QTLs for WBR (qWBR2 and qWBR5) showed similar chromosomal locations with GW2 and qSW5/GW5, which have been cloned previously to control the grain width and chalkiness (Huang et al., 2015; Gong et al., 2017).
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5.2
Rice
The cloning of Chalk5 and identification of other QTLs
By contrast, only a few QTLs responsible for grain chalkiness have been finely mapped and functionally characterized. Chalk5, the only cloned and functionally characterized chalkiness gene, was located on chromosome 5 and encodes a vacuolar Hþtranslocating pyrophosphatase. Li et al. (2014) found that Chalk5 expressed specially in endosperm is a positive regulator of grain white belly rate, and the ZS97 allele greatly increased chalkiness rate in near-isogenic lines and transgenic lines (Fig. 11.3B). Two SNPs in the promoter region of chalk5 repressed the expression level of Chalk5, resulting in a decrease of chalkiness. Sequencing of cultivated rice and genome-wide association analysis further revealed that these two SNPs are the main reason for the variation of chalkiness in indica. Besides chalk5, there are many starch synthesis genes and grain shape genes related to grain chalkiness. Loss of function of Flo5/SSIIIa, which participates in the elongation of short-chain starch, will influence the structure of starch, and T-DNA insertion of Flo5 displays a white core phenotype. It has been thought that there is a significant positive relation between grain width and chalkiness, and many cloned grain width genes do indeed influence grain chalkiness.
5.3
Molecular mechanisms of chalkiness formation
Due to the lack of cloned chalkiness-related genes and research of the molecular mechanism, the cause of chalkiness is poorly understood. As understood from existing research results, chalkiness is formed by the loose arrangement between starch granules and proteasome in endosperm during grain-filling stage and might be a result of different accumulation rates of nutrition parts due to the different transport channels of photosynthesis products. Ishimaru et al. (2009) tried to figure out the cause of chalkiness through heat stress and found that high temperature can induce the formation of chalk. They further proved that the abnormal loss of water of endosperm under high temperature was the main cause of white core and floury phenotype. The research of Li et al. (2014) demonstrated that elevated expression of Chalk5 increases chalkiness rate by disturbing the pH homeostasis in the endomembrane trafficking system in developing seeds, which affects the biogenesis of protein bodies and increases the small vesicle-like structures, leading to the formation of air spaces among endosperm storage substances and resulting in chalky grains (Fig. 11.3A and C).
6.
Genetic improvement of rice appearance quality
Development of new cultivars with improved appearance quality is important for rice breeding. The identification and cloning of appearance quality QTLs provide gene sources for marker-assisted selection (MAS) and genetic engineering. Fan et al. (2006) cloned the major grain length gene GS3 and developed a functional molecular marker, SF28, according to the C to A variation. Wang et al. (2012) utilized SF28 for the MAS of GS3, to pyramid the nonfunctional allele gs3 and gw8 into HJX74, a
Rice appearance quality
(A)
379
(B)
Golgi apparatus Nucleus
Prevacuolar compartment
Zhenshan 97
OHH+ OHH+ H + H2 O H2 O
Chalk5 Water loss
H94
(C)
Vacuole (proteosome) NIL(H94)
NIL(ZS97)
OX(–)
OX(+)
ZpZc(–)
ZpZc(+)
Figure 11.3 Chalk5 increase grain chalkiness in rice. (A) A proposed working model for Chalk5. Chalk5 accelerates the water loss of vacuole and induces the accumulation of small vesicle-like structures, leading to the formation of air spaces among endosperm storage substances, resulting in chalky grains. (B) Grain chalkiness of zhenshan 97, H94, NIL(ZS97), NIL(H94), and the two transformants in T2 progeny. OX(þ) and ZpZc(þ) indicate grains from T2 plants expressing the coding region of Chalk5 from Zhenshan 97 driven by the 35S promoter and the native promoter from Zhenshan 97, respectively. OX() and ZpZc() represent the corresponding transgene-negative segregants. (C) Scanning electron microscopy images of transverse sections from the endosperm bellies of mature seeds. Positions marked by rectangles in the middle bellies of the endosperm were analyzed by scanning electron microscopy. Scale bars: 500 mm (top), 10 mm (middle), 5 mm (bottom).
variety with short and wide grains. They successfully improved the grain shape of HJX74 and formed a similar grain shape to Basimati385, a variety with slender grain. MAS of superior appearance quality genes was also applied to hybrid rice breeding. QTL pyramiding of the GW7 and gs3 alleles has helped the development of new high-yielding indica hybrid rice varieties (for example, Taifengyou 55 and Taifengyou 208) with simultaneously improved yield and grain quality (Wang et al., 2015a,b). Unlike grain shape, little progress has been made in selecting against chalk over the years. Zhao et al. (2016) found that low chalk lines in the RIL population maintained their chalk classification across the nine environments. And introgression of five QTLs together almost eliminated grain chalkiness. CRISPR/Cas9 systems have been successfully used as efficient tools for genome editing in a variety of species. We can knock out negative regulators of grain quality efficiently to speed up the breeding process in rice. Li et al. (2016) used a CRISPR/ Cas9 system to edit four yield-related genes (Gn1a, DEP1, GS3, and IPA1) in Zhonghua 11 and obtained phenotypes similar to those of previously reported mutants, respectively. Another study targeted eight genes in rice (including GS3 and GW2) using a CRISPR/Cas9 multiplex genome editing system, and various phenotypes related to the editing genes were observed (Shen et al., 2017).
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7.
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Major issues and prospects in studies on rice appearance quality
Over the past decade, scientists have made impressive achievements in genomic studies on rice appearance quality. Those achievements can be summed up in three aspects: (1) the discovery of important genes for rice appearance quality; (2) the progress in molecular properties, expression and regulation of genes; (3) the application of MAS and transgenic technology. However, rice appearance quality is composed of many traits, and there were pervasive interactions among quality traits, between quality and yield, as well as between quality and environment. Additionally, the key scientific problem of rice appearance quality is not obvious to people. Those issues limit the improvement of rice appearance quality. Nonchalkiness is the primary goal in genetic improvement and basic research of rice appearance quality. Rice chalkiness is a trait conferring all the milling, appearance, eating, and cooking qualities. As the white rices of most varieties have approximately the same nutrition quality, the chalkiness tends to be the crucial problem need to be solved in rice quality improvement. The general high chalkiness rate of indica hybrid rice forms a common view about the negative correlation between rice quality and yield. However, few indica hybrid rice varieties are of both good quality and yield, which indicates that quality can coexist with yield if we can figure out their genetic mechanisms. Currently, the following measures are useful in reducing rice chalkiness: (1) properly improve grain length to width and break the adverse linkage between chalkiness QTLs and grain width QTLs; (2) take advantage of diverse natural germplasm resources and artificial mutant resources to identify and clone more chalkiness functional genes, and to ascertain the molecular, celled, physiological, and biochemical mechanism during chalkiness formation; (3) look for varieties with low chalkiness in heat stress and utilize their superior genes to solve the problem of chalkiness formation in high temperature. Sustained high temperatures after rice heading will greatly increase the proportion of chalky rice. In Asian rice growing areas, this climatic condition now occurs more often than it did in previous years, possibly due to global warming. Global warming will become a global issue in future agriculture. Thus, thorough and comprehensive genetic studies on rice quality are needed to face the challenges of future climate change.
References Bowles, D., 2012. Towards increased crop productivity and quality. Current Opinion in Biotechnology 23, 202e203. Cheng, F., Zhong, L., Wang, F., Zhang, G., 2005. Differences in cooking and eating properties between chalky and translucent parts in rice grains. Food Chemistry 90, 39e46. Duan, P., Xu, J., Zeng, D., Zhang, B., Geng, M., Zhang, G., Huang, K., Huang, L., Xu, R., Ge, S., 2017. Natural variation in the promoter of GSE5 contributes to grain size diversity in rice. Molecular Plant 10, 685e694.
Rice appearance quality
381
Fan, C., Xing, Y., Mao, H., Lu, T., Han, B., Xu, C., Li, X., Zhang, Q., 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. Theoretical and Applied Genetics 112, 1164e1171. Fitzgerald, M.A., McCouch, S.R., Hall, R.D., 2009. Not just a grain of rice: the quest for quality. Trends in Plant Science 14, 133e139. Furukawa, T., Maekawa, M., Oki, T., Suda, I., Iida, S., Shimada, H., Takamure, I., Kadowaki, K., 2007. The Rc and Rd genes are involved in proanthocyanidin synthesis in rice pericarp. The Plant Journal 49, 91e102. Gong, J., Jiashun, M., Yan, Z., Qiang, Z., Qi, F., Qilin, Z., Benyi, C., Junhui, X., Xuehui, H., Han, B., 2017. Dissecting the genetic basis of grain shape and chalkiness traits in hybrid rice using multiple collaborative populations. Molecular Plant 10, 1353e1356. Hu, J., Wang, Y., Fang, Y., Zeng, L., Xu, J., Yu, H., Shi, Z., Pan, J., Zhang, D., Kang, S., Zhu, L., Dong, G., Guo, L., Zeng, D., Zhang, G., Xie, L., Xiong, G., Li, J., Qian, Q., 2015. A rare allele of GS2 enhances grain size and grain yield in rice. Molecular Plant 8, 1455e1465. Huang, R., Jiang, L., Zheng, J., Wang, T., Wang, H., Huang, Y., Hong, Z., 2013. Genetic bases of rice grain shape: so many genes, so little known. Trends in Plant Science 18, 218e226. Huang, X., Yang, S., Gong, J., Zhao, Y., Feng, Q., Gong, H., Li, W., Zhan, Q., Cheng, B., Xia, J., 2015. Genomic analysis of hybrid rice varieties reveals numerous superior alleles that contribute to heterosis. Nature Communications 6, 6258. Ishimaru, T., Horigane, A.K., Ida, M., Iwasawa, N., San-oh, Y.A., Nakazono, M., Nishizawa, N.K., Masumura, T., Kondo, M., Yoshida, M., 2009. Formation of grain chalkiness and changes in water distribution in developing rice caryopses grown under high-temperature stress. Journal of Cereal Science 50, 166e174. Ishimaru, K., Hirotsu, N., Madoka, Y., Murakami, N., Hara, N., Onodera, H., Kashiwagi, T., Ujiie, K., Shimizu, B-i, Onishi, A., 2013. Loss of function of the IAA-glucose hydrolase gene TGW6 enhances rice grain weight and increases yield. Nature Genetics 45, 707e711. Lanning, S.B., Siebenmorgen, T.J., Counce, P.A., Ambardekar, A.A., Mauromoustakos, A., 2011. Extreme nighttime air temperatures in 2010 impact rice chalkiness and milling quality. Field Crops Research 124, 132e136. Li, Y., Fan, C., Xing, Y., Jiang, Y., Luo, L., Sun, L., Shao, D., Xu, C., Li, X., Xiao, J., He, Y., Zhang, Q., 2011. Natural variation in GS5 plays an important role in regulating grain size and yield in rice. Nature Genetics 43, 1266e1269. Li, Y., Fan, C., Xing, Y., Yun, P., Luo, L., Yan, B., Peng, B., Xie, W., Wang, G., Li, X., Xiao, J., Xu, C., He, Y., 2014. Chalk5 encodes a vacuolar H(þ)-translocating pyrophosphatase influencing grain chalkiness in rice. Nature Genetics 46, 398e404. Li, M., Li, X., Zhou, Z., Wu, P., Fang, M., Pan, X., Lin, Q., Luo, W., Wu, G., Li, H., 2016. Reassessment of the four yield-related genes Gn1a, DEP1, GS3, and IPA1 in rice using a CRISPR/Cas9 system. Frontiers in plant science 7, 377. Liu, J., Chen, J., Zheng, X., Wu, F., Lin, Q., Heng, Y., Tian, P., Cheng, Z., Yu, X., Zhou, K., 2017. GW5 acts in the brassinosteroid signalling pathway to regulate grain width and weight in rice. Nature Plants 3, 17043. Mao, H., Sun, S., Yao, J., Wang, C., Yu, S., Xu, C., Li, X., Zhang, Q., 2010. Linking differential domain functions of the GS3 protein to natural variation of grain size in rice. Proceedings of the National Academy of Sciences of the United States of America 107, 19579e19584. Oikawa, T., Maeda, H., Oguchi, T., Yamaguchi, T., Tanabe, N., Ebana, K., Yano, M., Ebitani, T., Izawa, T., 2015. The birth of a black rice gene and its local spread by introgression. The Plant Cell 27, 2401e2414.
382
Rice
Paine, J.A., Shipton, C.A., Chaggar, S., Howells, R.M., Kennedy, M.J., Vernon, G., Wright, S.Y., Hinchliffe, E., Adams, J.L., Silverstone, A.L., 2005. Improving the nutritional value of Golden Rice through increased pro-vitamin A content. Nature Biotechnology 23, 482e487. Peng, B., Wang, L., Fan, C., Jiang, G., Luo, L., Li, Y., He, Y., 2014. Comparative mapping of chalkiness components in rice using five populations across two environments. BMC Genetics 15, 49. Qi, P., Lin, Y.S., Song, X.J., Shen, J.B., Huang, W., Shan, J.X., Zhu, M.Z., Jiang, L., Gao, J.P., Lin, H.X., 2012. The novel quantitative trait locus GL3.1 controls rice grain size and yield by regulating Cyclin-T1;3. Cell Research 22, 1666e1680. Shen, L., Hua, Y., Fu, Y., Li, J., Liu, Q., Jiao, X., Xin, G., Wang, J., Wang, X., Yan, C., 2017. Rapid generation of genetic diversity by multiplex CRISPR/Cas9 genome editing in rice. Science China Life Sciences 1e10. 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. Nature Genetics 40, 1023e1028. Si, L., Chen, J., Huang, X., Gong, H., Luo, J., Hou, Q., Zhou, T., Lu, T., Zhu, J., Shangguan, Y., 2016. OsSPL13 controls grain size in cultivated rice. Nature Genetics 48, 447e456. Siebenmorgen, T.J., Grigg, B.C., Lanning, S.B., 2013. Impacts of preharvest factors during kernel development on rice quality and functionality. Annual Review of Food Science and Technology 4, 101e115. Singh, N., Sodhi, N., Kaur, M., Saxena, S., 2003. Physico-chemical, morphological, thermal, cooking and textural properties of chalky and translucent rice kernels. Food Chemistry 82, 433e439. Song, X.-J., Huang, W., Shi, M., Zhu, M.-Z., Lin, H.-X., 2007. A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase. Nature Genetics 39, 623e630. Sundaram, R.M., Vishnupriya, M.R., Biradar, S.K., Laha, G.S., Reddy, G.A., Rani, N.S., Sarma, N.P., Sonti, R.V., 2008. Marker assisted introgression of bacterial blight resistance in Samba Mahsuri, an elite indica rice variety. Euphytica 160, 411e422. Sweeney, M.T., Thomson, M.J., Pfeil, B.E., McCouch, S., 2006. Caught red-handed: Rc encodes a basic helix-loop-helix protein conditioning red pericarp in rice. The Plant Cell 18, 283e294. Takano-Kai, N., Jiang, H., Kubo, T., Sweeney, M., Matsumoto, T., Kanamori, H., Padhukasahasram, B., Bustamante, C., Yoshimura, A., Doi, K., McCouch, S., 2009. Evolutionary history of GS3, a gene conferring grain length in rice. Genetics 182, 1323e1334. Wang, E., Wang, J., Zhu, X., Hao, W., Wang, L., Li, Q., Zhang, L., He, W., Lu, B., Lin, H., 2008. Control of rice grain-filling and yield by a gene with a potential signature of domestication. Nature Genetics 40, 1370e1374. Wang, S., Wu, K., Yuan, Q., Liu, X., Liu, Z., Lin, X., Zeng, R., Zhu, H., Dong, G., Qian, Q., Zhang, G., Fu, X., 2012. Control of grain size, shape and quality by OsSPL16 in rice. Nature Genetics 44, 950e954. Wang, S., Li, S., Liu, Q., Wu, K., Zhang, J., Wang, S., Wang, Y., Chen, X., Zhang, Y., Gao, C., 2015a. The OsSPL16-GW7 regulatory module determines grain shape and simultaneously improves rice yield and grain quality. Nature Genetics 47, 949e954. Wang, Y., Xiong, G., Hu, J., Jiang, L., Yu, H., Xu, J., Fang, Y., Zeng, L., Xu, E., Xu, J., Ye, W., Meng, X., Liu, R., Chen, H., Jing, Y., Wang, Y., Zhu, X., Li, J., Qian, Q., 2015b. Copy number variation at the GL7 locus contributes to grain size diversity in rice. Nature Genetics 47, 944e948.
Rice appearance quality
383
Weng, J., Gu, S., Wan, X., Gao, H., Guo, T., Su, N., Lei, C., Zhang, X., Cheng, Z., Guo, X., Wang, J., Jiang, L., Zhai, H., Wan, J., 2008. Isolation and initial characterization of GW5, a major QTL associated with rice grain width and weight. Cell Research 18, 1199e1209. Xia, D., Zhou, H., Liu, R., Dan, W., Li, P., Wu, B., Chen, J., Wang, L., Gao, G., Zhang, Q., He, Y., 2018. GL3. 3, a Novel QTL Encoding a GSK3/SHAGGY-like Kinase, Epistatically Interacts with GS3 to Produce Extra-long Grains in Rice. Molecular plant 11, 754e756. Xu, C., Liu, Y., Li, Y., Xu, X., Xu, C., Li, X., Xiao, J., Zhang, Q., 2015. Differential expression of GS5 regulates grain size in rice. Journal of Experimental Botany 66, 2611e2623. Yan, W.G., Bao, J.S., 2014. Genes and QTLs for rice grain quality improvement. RiceGermplasm. Genetics and Improvement (InTech), pp. 239e278. Ye, X., Al-Babili, S., Kl€oti, A., Zhang, J., Lucca, P., Beyer, P., Potrykus, I., 2000. Engineering the provitamin A (b-carotene) biosynthetic pathway into (carotenoid-free) rice endosperm. Science 287, 303e305. Yun, P., Zhu, Y., Wu, B., Gao, G., Sun, P., Zhang Q,He, Y., 2016. Genetic mapping and confirmation of quantitative trait loci for grain chalkiness in rice. Molecular Breeding 36, 162. Zhao, X., Daygon, V.D., McNally, K.L., Hamilton, R.S., Xie, F., Reinke, R.F., Fitzgerald, M.A., 2016. Identification of stable QTLs causing chalk in rice grains in nine environments. Theoretical and Applied Genetics 129, 141e153. Zhou, H., Li, P., Xie, W., Hussain, S., Li, Y., Xia, D., Zhao, H., Sun, S., Chen, J., Ye, H., 2017. Genome-wide association analyses reveal the genetic basis of stigma exsertion in rice. Molecular Plant 10, 634e644. Zhu, Q., Yu, S., Zeng, D., Liu, H., Wang, H., Yang, Z., Xie, X., Shen, R., Tan, J., Li, H., 2017. Development of “purple endosperm rice” by engineering anthocyanin biosynthesis in the endosperm with a high-efficiency transgene stacking system. Molecular Plant 10, 918e929. Zuo, J., Li, J., 2014. Molecular genetic dissection of quantitative trait loci regulating rice grain size. Annual Review of Genetics 48, 99e118.