1/25/2019 International Plant & Animal Genome XXVII W900 / January 14, 2019 San Diego, CA, USA An Ethylene-Gibberellin Relay Co-opts “Deepwater rice” the Green Revolution Gene to Allow Rice Adaptation to Submergence Takeshi Kuroha (Tohoku Univ.) Susan McCouch (Cornell Univ.) Moto Ashikari (Nagoya Univ.) Deepwater rice field in Thailand Deepwater rice field in Thailand Deepwater response (Tank was filled to the brim at the first day) Water is essential for all living organisms, and plant lives around swamps and rivers has an advantage in terms of water availability NASA IRRI Excess water due to flooding has an adverse effect on the growth of most plants Non-Deepwater rice Deepwater rice Hattori et al 2009 Nature Dry season Hollow structure in the stem is used for air flow Snorkel Node Rainy season Internode Node There are plants that can adapt to periodic flooding Internode http://angkorvat.jp Shallow Deepwater 1 1/25/2019 Previous model of internode elongation in deepwater rice Genome wide association study (GWAS) on deepwater response Collaboration with Prof. Mochizuki (Kyushu Univ.) (Non-deepwater rice; Deepwater rice) Korea Deepwater rice and Dr. Ebana (NARO) Non-deepwater rice (1; 0) Submergence Submergence Bhutan (1; 0) China Total 68 varieties (5; 0) (28 non-deepwater and 40 deepwater rice) Nepal Ethylene Ethylene Taiwan (1; 0) India (1; 0) accumulation accumulation (9; 3) Philippines (4; 0) Vietnam OsEIL1a OsEIL1a (0; 8) Bangladesh (1; 20) Cambodia (1; 2) Myanmar Thailand SNORKEL1/ 2 SNORKEL1/2 Madagascar (2; 4) (0; 4) × (1; 0) TILS ? (Total internode length after one week submergence) Gibberellin accumulation Water level Drowning Internode elongation Air Submergence for 1 week Procedure of submergence treatment Togo field in Nagoya Univ. Outline 1. Identification of the gene for internode elongation under submergence in deepwater rice 2. Special function of Green revolution gene in deepwater rice to adapt periodical flooding Deepwater rice field in Thailand Explorations of regulatory factors for deepwater response Result of GWAS on deepwater response Hattori et al. Breeding Sci. 2008 Collaboration with Prof. Tamiya (Tohoku Univ./RIKEN) Nagai et al. Breeding Sci. 2012 and Prof. Yamasaki (Kobe Univ.) Biparental linkage analysis Deepwater response of 68 varieties for GWAS chromosomes Sub: Submergence for 1 week 1 2 3 4 5 6 7 8 9 10 11 12 Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Air Sub Genotyping with genetic makers 3 and mapping 1 VS m 4 12 2 1 T65 NIAS37 NIAS44 NIAS45 NIAS01 DW470 DW311 DW414 DW426 Bhadua C9285 T65 C9285 SNORKEL1/2 (non-deepwater rice) (deepwater rice) Ethylene response factor TILS values of each variety Manhattan plot for trait of TILS Hattori et al Nature 2009 Genome wide association study (GWAS) (Deepwater rice) based on haplotype association analysis C9285 chromosomes 7 120 6 1 2 3 4 5 6 7 8 9 10 11 12 100 ) 5 P 80 ( 4 Genotyping with 10 44k DNA microarray 3 60 log and mapping – ? 2 TILS(cm) 40 (Non-deepwater rice) ・・・ 1 T65 20 0 0 1 2 3 4 5 6 7 8 9 10 11 12 T65 C9285 DW314 DW478 DW483 DW487 DW316 DW313 DW417 DW466 DW489 DW318 DW397 DW471 DW311 DW456 DW457 DW375 DW369 NIAS44 NIAS45 NIAS21 NIAS42 NIAS15 NIAS53 NIAS06 NIAS39 NIAS46 NIAS08 NIAS31 NIAS05 NIAS09 Bhadua Various non-deepwater and deepwater rice in Asia NIAS38 Varieties (68 lines) Chromosome Kuroha et al Science 2018 2 1/25/2019 Strong peak on chromosome 1 Polymorphisms in SD1 are associate with deepwater response Collaboration with Prof. Tamiya (Tohoku Univ./RIKEN) 34.5 42.5 (Mb) Chr.1 and Prof. Yamasaki (Kobe Univ.) Nucleotide variation in SD1 gene of varieties in the GWAS Insertion Deletion Substitution GWAS 8 qTIL1C9285 Hattori et al 2008 The peak of GWAS on chr1 was Haplotype n ) 6 C9285 P T65 4 ( located inside of the QTL (qTIL1 ), T65 10 aus 23 4 which was previously detected by C9285 log indica 25 – other biparental linkage analysis 14 2 C9285 0 T65 C9285 Interaction between TILS, SD1 haplotypes, 36 38 40 42 (Mb) and existence of SNORKEL1/2 1 2 3 4 5 6 7 8 9 101112 (chr.) 1 2 3 4 5 6 7 8 9 101112 3.7 Mb 4.0 Mb SNORKEL1/2 DWH (Deepwater rice specific haplotype) × Linkage mapping 0.35 Mb ・17 polymorphisms are associated with internode NIL-12 NIL-1+12 elongation under submergence HindIII Indel Marker ・The DWH works in the presence of SNORKEL1/2 3379 3561N 32k 3227SSR High resolution linkage mapping to enhance internode elongation 32 kb was performed 169 kb 148 kb C9285 Kuroha et al Science 2018 Kuroha et al Science 2018 High-resolution linkage analysis of the QTL on chromosome 1 New model of internode elongation in deepwater rice Collaboration with Dr. Wu (NARO) Deepwater rice Chromosome 1 Chr. 1 2 3 4 5 6 7 8 9 101112 1 2 3 4 5 6 7 8 9 101112 1 2 3 4 5 6 7 8 9 101112 Submergence >1,000 recombination events Ethylene accumulation Indel SD1 5.5 kb SD106 SD103 32k 3561N ? T65 C9285 NIL-1 OsEIL1a T65 C9285 NIL-1 Os01g0883800 ? 15 6 * Os01g0883900 SNORKEL1/2 DWH SD1 * * 10 * 4 transcript transcript level SD1 (SEMI DWARF1) 5 2 SD1 ・Gene involved in modern breeding including the “Green Revolution” ・ 0 0 Loss-of-function of the gene leads semi-dwarf phenotype Relative 0 3 0 1 ・encodes a gibberellin biosynthesis enzyme, OsGA20ox2 After submergence (h) After ethylene treatment (h) Both submergence and ethylene induce SD1 expression WT sd1 Sasaki et al Nature 2002 in deepwater rice Internode elongation Kuroha et al Science 2018 Kuroha et al Science 2018 New model of internode elongation in deepwater rice Collaboration with Dr. Mitsuda, Dr. Ohme-Takagi, and Dr. Yanagisawa Deepwater rice submergence Submergence Outline Ethylene accumulation OsEIL1a fused with the repression domain inhibited SD1 induction under submergence 1. Identification of the gene for internode elongation ? OsEIL1a under submergence in deepwater rice Suppressor ? × SNORKEL1/2 DWH SD1 2. Special function of Green revolution gene Relative LUC activity in deepwater rice to adapt periodical flooding Reporter 0 50 100 LUC NBS MYC (bp) Effector OsEIL1a -3,551 +30 pSD1-3551LUC pSD1T65 * T65 -3,153 +30 OsEIL1a directly activates the pSD1-3153LUC SD1 promoter in rice protoplast pSD1C9285 * Deepwater rice field in Thailand Internode elongation Kuroha et al Science 2018 3 1/25/2019 New model of internode elongation in deepwater rice Origin of the DWH Collaboration with Dr. Sakakibara 0h 12h 24h Geographical distribution of DWH in wild and cultivated rice Deepwater rice GA1 GA4 O. rufipogon Submergence Bangladesh (n = 258) O. rufipogon Ethylene accumulation non-DWH DWH Standing variation OsEIL1a DWH In South/Southeast Asia non-DWH Active gibberellins, GA1 and GA4, increased under submergence Suppressor ? Country having × DWH-positive accessions ) 60 SNORKEL1/2 DWH SD1 Artificial selection m × c 10.4 ( in Bangladesh h t g O. sativa n 40 Bangladesh e l O. sativa e (n = 235) Gibberellin accumulation d o non-DWH n r e 20 t GA1 GA n 4 i l 1 ×1.0 DWH a 0 t c o m T 0 Deepwater rice 5 5 5 5 5 Promotion of 5 8 8 6 6 8 6 6 in Bangladesh 8 5 5 5 5 5 5 5 5 2 2 2 2 T T T T 6 8 8 6 6 8 8 internode elongation 6 9 9 9 9 2 2 T 2 2 T T T C 9 9 9 9 C C C C C C C Control DWH Control GA GA GA1 GA4 1 4 non-DWH 2 weeks from germination under air condition, 10-5 M gibberellin treatment with 10-6 M uniconazole Internode elongation Deepwater rice cultivation GA4 strongly induces internode elongation in deepwater rice Kuroha et al Science 2018 New model of internode elongation in deepwater rice Contribution of SD1 gene to rice breeding for two different purposes Non-deepwater rice Deepwater rice Amplification of an ethylene-gibberellin relay Submergence Submergence Internode elongation under submergence Green revolution Ethylene accumulation Ethylene accumulation sd1 mutation DWH SD1 SD1 SD1 Loss of function Gain of function OsEIL1a OsEIL1a Ethylene- gibberellin relay Lodging sensitive SNORKEL1/2 × Suppressor ? Suppressor ? Survive Lodging resistant × under deep water SD1 SNORKEL1/2 DWH SD1 Drown under deep water × Gibberellin accumulation Gibberellin accumulation GA1 GA4 Promotion of internode elongation Semi-dwarf rice Normal rice Deepwater rice Human selected two SD1 alleles with opposite function for agriculture Drowning Internode elongation Haplotype network of the SD1 Acknowledgement Tohoku Univ. /RIKEN AIP Haplotype network of the SD1 with O. sativa and O. rufipogon Gen Tamiya Nagoya Univ. Tohoku Univ. Kyushu Univ. Keisuke Nagai Shinjiro Yamaguchi Toshihiro Mochizuki aus Subpopulation based on Rico Gamuyao (present: Kyoto Univ.) Univ. of Tokyo genome wide SNP data (present: Johns Hopkins Univ. Kiyoshi Mashiguchi Shuichi Yanagisawa indica School of Medicine) Yoshiya Seto aus RIKEN CSRS/Nagoya Univ. Japonica Tomoyuki Furuta (present: Meiji Univ.) DWH aro Hitoshi Sakakibara indica Masanari Nakamori Kazuhiko Nishitani tropical japonica RIKEN CSRS temperate japonica Takuya Kitaoka Ryusuke Yokoyama Mikiko Kojima admixed Keita Adachi O. rufipogon Kobe Univ. Anzu Minami Cornell Univ. Masanori Yamasaki Yoshinao Mori Diane R. Wang NIAS Deepwater rice (present: Univ. at Buffalo) in O. sativa Kaworu Ebana indica Jianzhong Wu aus JST CREST JICA-JST SATREPS AIST O. rufipogon Admixed (C9285) the Japan Advanced Plant Science Network Masaru Ohme-Takagi The Canon Foundation The DWH was found only in deepwatar rice within O. sativa and MEXT/JSPS KAKENHI (present: Saitama Univ.) a part of O.