Generation of High-Amylose Rice through CRISPR/Cas9-Mediated Targeted Mutagenesis of Starch Branching Enzymes

[en] Cereals high in amylose content (AC) and resistant starch (RS) offer potential health benefits. Previous studies using chemical mutagenesis or RNA interference have demonstrated that starch branching enzyme (SBE) plays a major role in determining the fine structure and physical properties of starch. However, it remains a challenge to control starch branching in commercial lines. Here, we use CRISPR/Cas9 technology to generate targeted mutagenesis in SBEI and SBEIIb in rice. The frequencies of obtained homozygous or bi-allelic mutant lines with indels in SBEI and SBEIIb in T0 generation were from 26.7 to 40%. Mutations in the homozygous T0 lines stably transmitted to the T1 generation and those in the bi-allelic lines segregated in a Mendelian fashion. Transgene-free plants carrying only the frame-shifted mutagenesis were recovered in T1 generation following segregation. Whereas no obvious differences were observed between the sbeI mutants and wild type, sbeII mutants showed higher proportion of long chains presented in debranched amylopectin, significantly increased AC and RS content to as higher as 25.0% and 9.8%, respectively, and thus altered fine structure and nutritional properties of starch. Taken together, our results demonstrated for the first time the feasibility to create high-amylose rice through CRISPR/Cas9-mediated editing of SBEIIb.

Disciplines :

Biotechnology

Author, co-author :

Sun, Yongwei ; Université de Liège - ULiège > Doct. sc. agro. & ingé. biol. (Paysage)

Xia, Lanqin

Francis, Frédéric ; Université de Liège > Agronomie, Bio-ingénierie et Chimie (AgroBioChem) > Gestion durable des bio-agresseurs

Jiao, Guiai

Liu, Zupei

Zhang, Xin

Li, Jingying

Guo, Xiuping

Du, Wenming

Du, Jinlu

Zhao, Yunde

Language :

English

Title :

Generation of High-Amylose Rice through CRISPR/Cas9-Mediated Targeted Mutagenesis of Starch Branching Enzymes

Publication date :

March 2017

Journal title :

Frontiers in Plant Science

eISSN :

1664-462X

Publisher :

Frontiers Research Foundation, Lausanne, Switzerland

Volume :

Pages :

298

Peer reviewed :

Peer Reviewed verified by ORBi

Funding text :

This work is partly funded by the Ministry of Science and Technology of China (grant no. 2016YFD0100500) and the Ministry of Agriculture of China (grant no. 2016ZX08010-003). YS is supported by a GSCAAS-ULg Joint Ph.D. Program.

Available on ORBi :

since 20 July 2017

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Bibliography

AACC (1995). Approved Methods of the American Association of Cereal Chemists, 9th Edn. St. Paul, MN: AACC.
Asaoka, M., Okuno, K., Sugimoto, Y., Yano, M., Omura, T., and Fuwa, H. (1986). Characterization of endosperm starch from high-amylose mutants of rice (Oryza sativa L.). Starch 38, 114–117. doi: 10.1002/star.198603 80403
Asp, N.-G. (1992). Resistant starch: proceedings from the 2nd plenary meeting of EURESTA: European Flair Concerted Action no. 11 (COST 911): physiological implications of the consumption of resistant starch in man. Eur. J. Clin. Nutr. 46, S1–S148. doi: 10.1016/0924-2244(92)90153-n
Ball, S. G., and Morell, M. K. (2003). From bacterial glycogen to starch: understanding the biogenesis of the plant starch granule. Annu. Rev. Plant Biol. 54, 207–233. doi: 10.1146/annurev.arplant.54.031902. 134927
Bhattacharyya, M. K., Smith, A. M., Ellis, T. N., Hedley, C., and Martin, C. (1990). The wrinkled-seed character of pea described by Mendel is caused by a transposon-like insertion in a gene encoding starch-branching enzyme. Cell 60, 115–122. doi: 10.1016/0092-8674(90)90721-P
Blauth, S. L., Kim, K.-N., Klucinec, J., Shannon, J. C., Thompson, D., and Guiltinan, M. (2002). Identification of mutator insertional mutants of starch-branching enzyme 1 (sbe1) in Zea mays L. Plant Mol. Biol. 48, 287–297. doi: 10.1023/A:1013335217744
Blauth, S. L., Yao, Y., Klucinec, J. D., Shannon, J. C., Thompson, D. B., and Guiltinan, M. J. (2001). Identification of Mutator insertional mutants of starch-branching enzyme 2a in corn. Plant Physiol. 125, 1396–1405. doi: 10.1104/pp. 125.3.1396
Boyer, C. D., and Preiss, J. (1978). Multiple forms of (1→4)-α-D-glucan,(1→4)-α-D-glucan-6-glycosyl transferase from developing Zea mays L. kernels. Carbohydr. Res. 61, 321–334. doi: 10.1016/S0008-6215(00) 84492-4
Butardo, V. Jr., Fitzgerald, M. A., Rahman, S., and Gidley, M. J. (2008). “Efforts to capture high amylose in rice,” in Proceedings of the AACC International Annual Meeting 2008 (Eagan, MN: American Association of Cereal Chemists), A15–A15.
Butardo, V. M., Fitzgerald, M. A., Bird, A. R., Gidley, M. J., Flanagan, B. M., Larroque, O., et al. (2011). Impact of down-regulation of starch branching enzyme IIb in rice by artificial microRNA-and hairpin RNA-mediated RNA silencing. J. Exp. Bot. 62, 4927–4941. doi: 10.1093/jxb/err188
Carciofi, M., Blennow, A., Jensen, S. L., Shaik, S. S., Henriksen, A., Buléon, A., et al. (2012). Concerted suppression of all starch branching enzyme genes in barley produces amylose-only starch granules. BMC Plant Biol. 12:223. doi: 10.1186/1471-2229-12-223
Chen, L., Magliano, D. J., and Zimmet, P. Z. (2012). The worldwide epidemiology of type 2 diabetes mellitus—present and future perspectives. Nat. Rev. Endocrinol. 8, 228–236. doi: 10.1038/nrendo.2011.183
Cong, L., Ran, F. A., Cox, D., Lin, S., Barretto, R., Habib, N., et al. (2013). Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819–823. doi: 10.1126/science.1231143
Dumez, S., Wattebled, F., Dauvillee, D., Delvalle, D., Planchot, V., Ball, S. G., et al. (2006). Mutants of Arabidopsis lacking starch branching enzyme II substitute plastidial starch synthesis by cytoplasmic maltose accumulation. Plant Cell 18, 2694–2709. doi: 10.1105/tpc.105.037671
Endo, M., Mikami, M., and Toki, S. (2016). Biallelic gene targeting in rice. Plant Physiol. 170, 667–677. doi: 10.1104/pp.15.01663
Englyst, H. N., Kingman, S., and Cummings, J. (1992). Classification and measurement of nutritionally important starch fractions. Eur. J. Clin. Nutr. 46, S33–S50.
Feng, Z., Mao, Y., Xu, N., Zhang, B., Wei, P., Yang, D. L., et al. (2014). Multigeneration analysis reveals the inheritance, specificity, and patterns of CRISPR/Cas-induced gene modifications in Arabidopsis. Proc. Natl. Acad. Sci. U.S.A. 111, 4632–4637. doi: 10.1073/pnas.1400822111
Food and Agriculture Organization [FAO] (2004). International Year of Rice (Online). Available at: http://www.fao.org/rice2004/en/rice-us.htm (accessed March 13, 2006).
Gao, X., Chen, J., Dai, X., Zhang, D., and Zhao, Y. (2016). An effective strategy for reliably isolating heritable and Cas9-free Arabidopsis mutants generated by CRISPR/Cas9-mediated genome editing. Plant Physiol. 171, 1794–1800. doi: 10.1104/pp.16.00663
Gao, Y., and Zhao, Y. (2014). Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing. J. Integr. Plant Biol. 56, 343–349. doi: 10.1111/jipb.12152
Garwood, D. L., Shannon, J., and Creech, R. (1976). Starches of endosperms possessing different alleles at the amylose-extender locus in Zea mays L.[maize]. Cereal Chem. 53, 355–364.
Giacco, R., Clemente, G., Brighenti, F., Mancini, M., D’avanzo, A., Coppola, S., et al. (1998). Metabolic effects of resistant starch in patients with type 2 diabetes. Diabetes Nutr. Metab. 11, 330–335.
Hanashiro, I., Abe, J.-I., and Hizukuri, S. (1996). A periodic distribution of the chain length of amylopectin as revealed by high-performance anion-exchange chromatography. Carbohydr. Res. 283, 151–159. doi: 10.1016/0008-6215(95) 00408-4
Hiei, Y., Ohta, S., Komari, T., and Kumashiro, T. (1994). Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J. 6, 271–282. doi: 10.1046/j.1365-313X. 1994.6020271.x
Hoebler, C., Karinthi, A., Chiron, H., Champ, M., and Barry, J. (1999). Bioavailability of starch in bread rich in amylose: metabolic responses in healthy subjects and starch structure. Eur. J. Clin. Nutr. 53, 360–366. doi: 10.1038/sj. ejcn.1600718
Itoh, K., Ozaki, H., Okada, K., Hori, H., Takeda, Y., and Mitsui, T. (2003). Introduction of Wx transgene into rice wx mutants leads to both high-and low-amylose rice. Plant Cell Physiol. 44, 473–480. doi: 10.1093/pcp/pcg068
Jane, J., Chen, Y., Lee, L., Mcpherson, A., Wong, K., Radosavljevic, M., et al. (1999). Effects of amylopectin branch chain length and amylose content on the gelatinization and pasting properties of starch 1. Cereal Chem. 76, 629–637. doi: 10.1094/CCHEM.1999.76.5.629
Jiang, H., Lio, J., Blanco, M., Campbell, M., and Jane, J. L. (2010). Resistant-starch formation in high-amylose maize starch during kernel development. J. Agric. Food Chem. 58, 8043–8047. doi: 10.1021/jf1 01056y
Jinek, M., Chylinski, K., Fonfara, I., Hauer, M., Doudna, J. A., and Charpentier, E. (2012). A programmable dual-RNA–guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816–821. doi: 10.1126/science. 1225829
Jobling, S. A., Schwall, G. P., Westcott, R. J., Sidebottom, C. M., Debet, M., Gidley, M. J., et al. (1999). A minor form of starch branching enzyme in potato (Solanum tuberosum L.) tubers has a major effect on starch structure: cloning and characterisation of multiple forms of SBE A. Plant J. 18, 163–171. doi: 10.1046/j.1365-313X.1999.00441.x
Jones, H. D. (2015). Regulatory uncertainty over genome editing. Nat. Plants 1, 14011. doi: 10.1038/nplants.2014.11
Kim, K.-S., Hwang, H.-G., Kang, H.-J., Hwang, I.-K., Lee, Y.-T., and Choi, H.-C. (2005). Ultrastructure of individual and compound starch granules in isolation preparation from a high-quality, low-amylose rice, ilpumbyeo, and its mutant, G2, a high-dietary fiber, high-amylose rice. J. Agric. Food Chem. 53, 8745–8751. doi: 10.1021/jf051194a
Li, C., and Gilbert, R. G. (2016). Progress in controlling starch structure by modifying starch-branching enzymes. Planta 243, 13–22. doi: 10.1007/s00425-015-2421-2
Li, J., Sun, Y., Du, J., Zhao, Y., and Xia, L. (2016). Generation of targeted point mutations in rice by a modified CRISPR/Cas9 system. Mol. Plant doi: 10.1016/j.molp.2016.12.001 [Epub ahead of print].
Li, M., Li, X., Zhou, Z., Wu, P., Fang, M., Pan, X., et al. (2016). Reassessment of the four yield-related Genes Gn1a, DEP1, GS3, and IPA1 in rice using a CRISPR/Cas9 system. Front. Plant Sci. 7:377. doi: 10.3389/fpls.2016. 00377
Li, Z., Liu, Z.-B., Xing, A., Moon, B. P., Koellhoffer, J. P., Huang, L., et al. (2015). Cas9-guide RNA directed genome editing in soybean. Plant Physiol. 169, 960–970. doi: 10.1104/pp.15.00783
Ma, X., Zhang, Q., Zhu, Q., Liu, W., Chen, Y., Qiu, R., et al. (2015). A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants. Mol. Plant 8, 1274–1284. doi: 10.1016/j.molp.2015. 04.007
McCleary, B. V., McNally, M., and Rossiter, P. (2002). Measurement of resistant starch by enzymatic digestion in starch and selected plant materials: collaborative study. J. AOAC Int. 85, 1103–1111.
Mizuno, K., Kawasaki, T., Shimada, H., Satoh, H., Kobayashi, E., Okumura, S., et al. (1993). Alteration of the structural properties of starch components by the lack of an isoform of starch branching enzyme in rice seeds. J. Biol. Chem. 268, 19084–19091.
Mizuno, K., Kimura, K., Arai, Y., Kawasaki, T., Shimada, H., and Baba, T. (1992). Starch branching enzymes from immature rice seeds. J. Biochem. 112, 643–651. doi: 10.1093/oxfordjournals.jbchem.a123953
Morell, M. K., Konik-Rose, C., Ahmed, R., Li, Z., and Rahman, S. (2004). Synthesis of resistant starches in plants. J. AOAC Int. 87, 740–748.
Morell, M. K., and Myers, A. M. (2005). Towards the rational design of cereal starches. Curr. Opin. Plant Biol. 8, 204–210. doi: 10.1016/j.pbi.2005.01.009
Nakamura, Y. (2002). Towards a better understanding of the metabolic system for amylopectin biosynthesis in plants: rice endosperm as a model tissue. Plant Cell Physiol. 43, 718–725. doi: 10.1093/pcp/pcf091
Nakamura, Y., Takeichi, T., Kawaguchi, K., and Yamanouchi, H. (1992). Purification of two forms of starch branching enzyme (Q-enzyme) from developing rice endosperm. Physiol. Plant 84, 329–335. doi: 10.1111/j.1399-3054.1992.tb04672.x
Nishi, A., Nakamura, Y., Tanaka, N., and Satoh, H. (2001). Biochemical and genetic analysis of the effects of amylose-extender mutation in rice endosperm. Plant Physiol. 127, 459–472. doi: 10.1104/pp.010127
Ohdan, T., Francisco, P. B., Sawada, T., Hirose, T., Terao, T., Satoh, H., et al. (2005). Expression profiling of genes involved in starch synthesis in sink and source organs of rice. J. Exp. Bot. 56, 3229–3244. doi: 10.1093/jxb/eri292
Ossowski, S., Schwab, R., and Weigel, D. (2008). Gene silencing in plants using artificial microRNAs and other small RNAs. Plant J. 53, 674–690. doi: 10.1111/j.1365-313X.2007.03328.x
Paul, J. W., III., and Qi, Y. (2016). CRISPR/Cas9 for plant genome editing: accomplishments, problems and prospects. Plant Cell Rep. 35, 1417–1427. doi: 10.1007/s00299-016-1985-z
Puchta, H., and Fauser, F. (2014). Synthetic nucleases for genome engineering in plants: prospects for a bright future. Plant J. 78, 727–741. doi: 10.1111/tpj. 12338
Rahman, S., Bird, A., Regina, A., Li, Z., Ral, J. P., Mcmaugh, S., et al. (2007). Resistant starch in cereals: exploiting genetic engineering and genetic variation. J. Cereal Sci. 46, 251–260. doi: 10.1016/j.jcs.2007.05.001
Regina, A., Bird, A., Topping, D., Bowden, S., Freeman, J., Barsby, T., et al. (2006). High-amylose wheat generated by RNA interference improves indices of large-bowel health in rats. Proc. Natl. Acad. Sci. U.S.A. 103, 3546–3551. doi: 10.1073/pnas.0510737103
Regina, A., Kosar-Hashemi, B., Li, Z., Rampling, L., Cmiel, M., Gianibelli, M. C., et al. (2004). Multiple isoforms of starch branching enzyme-I in wheat: lack of the major SBE-I isoform does not alter starch phenotype. Funct. Plant Biol. 31, 591–601. doi: 10.1071/FP03193
Regina, A., Kosar-Hashemi, B., Ling, S., Li, Z., Rahman, S., and Morell, M. (2010). Control of starch branching in barley defined through differential RNAi suppression of starch branching enzyme IIa and IIb. J. Exp. Bot. 61, 1469–1482. doi: 10.1093/jxb/erq011
Regmi, P. R., Metzler-Zebeli, B. U., Gänzle, M. G., Van Kempen, T. A., and Zijlstra, R. T. (2011). Starch with high amylose content and low in vitro digestibility increases intestinal nutrient flow and microbial fermentation and selectively promotes bifidobacteria in pigs. J. Nutr. 141, 1273–1280. doi: 10.3945/jn.111. 140509
Safford, R., Jobling, S. A., Sidebottom, C. M., Westcott, R. J., Cooke, D., Tober, K. J., et al. (1998). Consequences of antisense RNA inhibition of starch branching enzyme activity on properties of potato starch. Carbohydr. Polym. 35, 155–168. doi: 10.1016/S0144-8617(97)00249-X
Satoh, H., Nishi, A., Yamashita, K., Takemoto, Y., Tanaka, Y., Hosaka, Y., et al. (2003). Starch-branching enzyme I-deficient mutation specifically affects the structure and properties of starch in rice endosperm. Plant Physiol. 133, 1111–1121. doi: 10.1104/pp.103.021527
Schwall, G. P., Safford, R., Westcott, R. J., Jeffcoat, R., Tayal, A., Shi, Y.-C., et al. (2000). Production of very-high-amylose potato starch by inhibition of SBE A and B. Nat. Biotechnol. 18, 551–554. doi: 10.1038/75427
Shan, Q., Wang, Y., Li, J., Zhang, Y., Chen, K., Liang, Z., et al. (2013). Targeted genome modification of crop plants using a CRISPR-Cas system. Nat. Biotechnol. 31, 686–688. doi: 10.1038/nbt.2650
Shan, Q., Zhang, Y., Chen, K., Zhang, K., and Gao, C. (2015). Creation of fragrant rice by targeted knockout of the OsBADH2 gene using TALEN technology. Plant Biotechnol. J. 13, 791–800. doi: 10.1111/pbi.12312
Shi, J., Gao, H., Wang, H., Lafitte, H. R., Archibald, R. L., Yang, M., et al. (2016). ARGOS8 variants generated by CRISPR-Cas9 improve maize grain yield under field drought stress conditions. Plant Biotechnol. J. 15, 207–216. doi: 10.1111/pbi.12603
Shu, X., Jiao, G., Fitzgerald, M. A., Yang, C., Shu, Q., and Wu, D. (2006). Starch structure and digestibility of rice high in resistant starch. Starch 58, 411–417. doi: 10.1002/star.200600501
Stinard, P. S., Robertson, D. S., and Schnable, P. S. (1993). Genetic isolation, cloning, and analysis of a mutator-induced, dominant antimorph of the maize amylose extender1 locus. Plant Cell 5, 1555–1566. doi: 10.1105/tpc.5.11.1555
Sun, Y., Li, J., and Xia, L. (2016a). Precise genome modification via sequence-specific nucleases-mediated gene targeting for crop improvement. Front. Plant Sci. 7:1928. doi: 10.3389/fpls.2016.01928
Sun, Y., Zhang, X., Wu, C., He, Y., Ma, Y., Hou, H., et al. (2016b). Engineering herbicide-resistant rice plants through CRISPR/Cas9-mediated homologous recombination of acetolactate synthase. Mol. Plant 9, 628–631. doi: 10.1016/j. molp.2016.01.001
Svitashev, S., Young, J. K., Schwartz, C., Gao, H., Falco, S. C., and Cigan, A. M. (2015). Targeted mutagenesis, precise gene editing, and site-specific gene insertion in maize using Cas9 and guide RNA. Plant Physiol. 169, 931–945. doi: 10.1104/pp.15.00793
Syahariza, Z., Sar, S., Hasjim, J., Tizzotti, M. J., and Gilbert, R. G. (2013). The importance of amylose and amylopectin fine structures for starch digestibility in cooked rice grains. Food Chem. 136, 742–749. doi: 10.1016/j.foodchem.2012. 08.053
Thompson, D. (2000). On the non-random nature of amylopectin branching. Carbohydr. Polym. 43, 223–239. doi: 10.1016/S0144-8617(00)00150-8
Unwin, N., Whiting, D., and Guariguata, L. (2013). International Diabetes Federation. IDF Diabetes Atlas. Brussels: International Diabetes Federation, 2012.
Vonk, R. J., Hagedoorn, R. E., De Graaff, R., Elzinga, H., Tabak, S., Yang, Y.-X., et al. (2000). Digestion of so-called resistant starch sources in the human small intestine. Am. J. Clin. Nutr. 72, 432–438.
Voytas, D. F., and Gao, C. (2014). Precision genome engineering and agriculture: opportunities and regulatory challenges. PLoS Biol. 12:e1001877. doi: 10.1371/journal.pbio.1001877
Wang, T. L., Bogracheva, T. Y., and Hedley, C. L. (1998). Starch: as simple as A. B, C? J. Exp. Bot. 49, 481–502. doi: 10.1093/jxb/49.320.481
Wang, Z.-P., Xing, H.-L., Dong, L., Zhang, H.-Y., Han, C.-Y., Wang, X.-C., et al. (2015). Egg cell-specific promoter-controlled CRISPR/Cas9 efficiently generates homozygous mutants for multiple target genes in Arabidopsis in a single generation. Genome Biol. 16, 144. doi: 10.1186/s13059-015-0715-0
Warthmann, N., Chen, H., Ossowski, S., Weigel, D., and Hervé, P. (2008). Highly specific gene silencing by artificial miRNAs in rice. PLoS ONE 3:e1829. doi: 10.1371/journal.pone.0001829
Wei, C., Qin, F., Zhou, W., Chen, Y., Xu, B., Wang, Y., et al. (2010). Formation of semi-compound C-type starch granule in high-amylose rice developed by antisense RNA inhibition of starch-branching enzyme. J. Agric. Food Chem. 58, 11097–11104. doi: 10.1021/jf1024533
Xie, K., Minkenberg, B., and Yang, Y. (2015). Boosting CRISPR/Cas9 multiplex editing capability with the endogenous tRNA-processing system. Proc. Natl. Acad. Sci. U.S.A. 112, 3570–3575. doi: 10.1073/pnas.1420294112
Yamakawa, H., Hirose, T., Kuroda, M., and Yamaguchi, T. (2007). Comprehensive expression profiling of rice grain filling-related genes under high temperature using DNA microarray. Plant Physiol. 144, 258–277. doi: 10.1104/pp.107. 098665
Yamanouchi, H., and Nakamura, Y. (1992). Organ specificity of isoforms of starch branching enzyme (Q-enzyme) in rice. Plant Cell Physiol. 33, 985–991. doi: 10.1111/j.1744-7909.2008.00714.x
Yano, M., Okuno, K., Kawakami, J., Satoh, H., and Omura, T. (1985). High amylose mutants of rice, Oryza sativa L. Theor. Appl. Genet. 69, 253–257. doi: 10.1007/BF00662436
Yao, Y., Thompson, D. B., and Guiltinan, M. J. (2004). Maize starch-branching enzyme isoforms and amylopectin structure. In the absence of starch-branching enzyme IIb, the further absence of starch-branching enzyme Ia leads to increased branching. Plant Physiol. 136, 3515–3523. doi: 10.1104/pp.104.043315
Zhou, H., Liu, B., Weeks, D. P., Spalding, M. H., and Yang, B. (2014). Large chromosomal deletions and heritable small genetic changes induced by CRISPR/Cas9 in rice. Nucleic Acids Res. 42, 10903–10914. doi: 10.1093/nar/gku806
Zhu, L., Gu, M., Meng, X., Cheung, S. C., Yu, H., Huang, J., et al. (2012). High-amylose rice improves indices of animal health in normal and diabetic rats. Plant Biotechnol. J. 10, 353–362. doi: 10.1111/j.1467-7652.2011.00667.x