Welcome to Acta Agronomica Sinica,

Acta Agronomica Sinica ›› 2024, Vol. 50 ›› Issue (6): 1514-1524.doi: 10.3724/SP.J.1006.2024.34154

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Preliminary study on the regulation of cassava plant type by MeLAZY1c gene

WANG Jia-Xiang1(), YU Xue-Ting1, LI Meng-Tao1, MAI Wei-Tao1, CHEN Xin2,3,*(), WANG Wen-Quan1,*()   

  1. 1College of Tropical Agriculture and Forestry / Sanya Nanfan Research Institute, Hainan University, Sanya 572000, Hainan, China
    2Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences / Hainan Key Laboratory of Conservation and Utilization of Tropical Agricultural Biological Resources, Hainan Institute of Tropical Agricultural Resources, Haikou 571101, Hainan, China
    3Sanya Research Institute, Chinese Academy of Tropical Agricultural Sciences, Sanya 572000, Hainan, China
  • Received:2023-09-12 Accepted:2024-01-12 Online:2024-06-12 Published:2024-02-09
  • Contact: * E-mail: chenxin@itbb.org.cn;E-mail: wangwenquan@itbb.org.cn
  • Supported by:
    National Natural Science Foundation of China NSFC-CG Joint Fund Key Project(3181101517);National Key Research and Development Program of China(2018YFD1000500);Innovation Research Project for Graduate Students in Hainan Province(Qhys2022-82)

Abstract:

The IGT gene family is involved in the regulation of crop plant type, and LAZY belongs to the IGT subfamily. By comparing the amino acid sequences of six Arabidopsis LAZY members as ‘seeds’ in the cassava genome, a total of 8 LAZY genes were identified in cassava, among which MeLAZY1c is highly homologous to AtLAZY1, which regulates branching angles. Based on this, MeLAZY1c had the highest transcription level in the stem by qRT-PCR and pMeLAZY1c was deeply stained in the vascular bundle by GUS staining, using MeLAZY1c as the experimental materials in this study. Eight photoresponsive/regulatory elements were found in the MeLAZY1c promoter, and we subsequently found that darkness significantly inhibited the relative expression level of MeLAZY1c. At the same time, gene editing of MeLAZY1c was performed and 19 homozygous edited lineages. The phenotype of MeLAZY1c mutants was observed after seedling transplantation, compared to the SC8 wild-type, the main stems of MeLAZY1c mutants had crawing growth, and the stem skin cells at the curved part were distorted and deformed with different sizes. The number of cells at 1 mm near the ground was about 1.5 times that of cells at the far ground. In conclusion, these results indicates that MeLAZY1c plays an important role in the establishment of upright/creeping growth of cassava.

Key words: cassava, plant type regulation, LAZY, light response, crawling, gene editing

Table 1

Primers used in the study"

引物名称
Primer name
引物序列
Primer sequence (5°-3°)
用途
Purpose
MeLAZY1c-cas9-F GATTATAGCAGAAGAGAATTCTGG 基因编辑
CRISPR-Cas9
MeLAZY1c-cas9-R AAACCCAGAATTCTCTTCTGCTAT
Tubulin-F GTGGAGGAACTGGTTCTGGA 实时荧光定量PCR
RT-qPCR
Tubulin-R TGCACTCATCTGCATTCTCC
MeLAZY1c-QPCR-F ATGCTGAAGAAAAAAATGTCCCA 实时荧光定量PCR
RT-qPCR
MeLAZY1c-QPCR-R TTCTTTTGGGGTTTATCTGCCTT
MeLAZY1c Hi-TOM-F ATAGAACATCCCTTGGCGAGC Hi-TOM 测序
Hi-TOM sequencing
MeLAZY1c Hi-TOM-R GCAGCAGAGCTCCTAGAAGAA
MeLAZY1c-Psl1p-F ACGCGTCGACATGAAGTTACTAGGTTGGATGC 亚细胞定位
Subcellular localization
MeLAZY1c-Psl1p-R CGGGATCCCAGCTCCAACACAAGGTAGT
MeLAZY1c-GUS-F AATTCGAGCTCGGTACCCGGGGATCCCCACAAGGAACATCAACTCAA GUS 载体连接
GUS carrier connection
MeLAZY1c-GUS-R CTCCTTTACTAGTCAGATCTACCATGGTTGACCAATAGCCTGTGTGATA

Fig. 1

LAZY family evolutionary tree and MeLAZY expression level A: LAZY family evolutionary tree; B: MeLAZY expression level in heat map."

Fig. 2

Transcription levels of MeLAZY1c in tissues of different varieties Different lowercase letters represent significant differences at P < 0.05."

Fig. 3

GUS staining of different tissues in pMeLAZY1c transgenic Arabidopsis thaliana A, E: cotyledons, axillary buds, and petioles; B, F: petiole and hypocotyl; C, G: hypocotyl; D, H: root and root tip. Bar: 100 μm in A-D, 200 μm in E-F, 100 μm in G-H."

Fig. 4

Cis-acting elements of MeLAZY1c promoter and detection of expression levels under light treatments during different periods A: distribution diagram of cis-acting elements of MeLAZY1c promoter; B: detection of expression levels of MeLAZY1c under light treatment at different stages. Different lowercase letters represent significant differences at P < 0.05."

Fig. 5

Subcellular localization of MeLAZY1c Bar: 30 μm."

Fig. 6

Editing types and quantity statistics of melazy1c mutants A: the editing type of melazy1c mutant by Hi-Tom high-throughput sequencing; B: statistics on the number of editing types of cassava melazy1c mutant."

Fig. 7

Phenotypic comparison between SC8 and melazy1c mutants A: phenotypic observation of SC8 and melazy1c mutants; B: the angle between petiole and main stem of melazy1c mutant and SC8; C: melazy1c mutant bending height. * represent significant difference (***, P < 0.001; ****, P < 0.0001)."

Fig. 8

Morphological observation of petiole cells in SC8 and melazy1c mutant"

Fig. 9

Observation on the morphology of stem epidermal cells in SC8 and melazy1c mutants A: cross section of SC8 main stem; B: cross section of the main stem of the melazy1c mutant; C: the enlarged cross-sectional view of SC8 main stem; D: the enlarged cross-sectional view of the main stem of the melazy1c mutant; E, G: the enlarged view of the near/far ground longitudinal section of SC8 main stem; F, H: melazy1c mutant curved near/far ground longitudinal section enlarged view; I: SC8 longitudinal cut; J: melazy1c mutant is longitudinally cut; K-L: statistics of the number of proximal/distal cells at 1 mm."

[1] 严华兵, 叶剑秋, 李开绵. 中国木薯育种研究进展. 中国农学通报, 2015, 31(15): 63-70.
doi: 10.11924/j.issn.1000-6850.casb14110159
Yan H B, Ye J Q, Li K M. Research progress in cassava breeding in China. Chin Agric Sci Bull, 2015, 31(15): 63-70. (in Chinese with English abstract)
[2] 蒋和平, 倪印峰, 朱福守. 中国木薯产业发展模式及对策建议. 农业展望, 2014, 10(8): 41-48.
Jiang H P, Ni Y F, Zhu F S. Development model and countermeasures for China’s cassava industry. Agric Outlook, 2014, 10(8): 41-48. (in Chinese with English abstract)
[3] 李军, 田益农, 盘欢, 罗燕春, 郑华. 木薯品种桂热4号的选育及栽培要点. 南方农业学报, 2014, 45: 1183-1187.
Li J, Tian Y N, Pan H, Luo Y C, Zheng H. Breeding and cultivation key points of cassava variety Guire 4. J Southern Agric, 2014, 45: 1183-1187. (in Chinese with English abstract)
[4] 李旭娟, 李纯佳, 徐超华, 刘洪博, 吴转娣, 林秀琴. 甘蔗MOC1基因(ScMOC1)的克隆与表达分析. 植物遗传资源学报, 2017, 18: 734-746.
doi: 10.13430/j.cnki.jpgr.2017.04.017
Li X J, Li C J, Xu C H, Liu H B, Wu Z D, Lin X Q. Cloning and expression analysis of sugarcane MOC1 gene (ScMOC1). J Plant Genet Resour, 2017, 18: 734-746 (in Chinese with English abstract).
[5] Li X, Qian Q, Fu Z. Control of tillering in rice. Nature, 2003, 422: 618-621.
[6] Doebley J, Stec A, Hubbard L. The evolution of apical dominance in maize. Nature, 1997, 386: 485-488.
[7] Yu B S, Lin Z W, Li H X, Li X J, Li J Y, Wang Y H, Zhang X, Zhu Z F, Zhai W X, Wang X K, Xie D X, Sun C Q. TAC1, a major quantitative trait locus controlling tiller angle in rice. Plant J, 2007, 52: 891-898.
doi: 10.1111/j.1365-313X.2007.03284.x pmid: 17908158
[8] Dardick C, Callahan A, Horn R, Ruiz K B, Zhebentyayeva T, Hollender C, Whitaker M, Abbott A, Scorza R. PpeTAC1 promotes the horizontal growth of branches in peach trees and is a member of a functionally conserved family found in diverse plants species. Plant J, 2013, 75: 618-630.
[9] Jin J, Huang W, Gao J P, Yang J, Shi M, Zhu M Z, Luo D, Lin H X. Genetic control of rice plant architecture under domestication. Nat Genet, 2008, 40: 1365-1369.
doi: 10.1038/ng.247 pmid: 18820696
[10] Tan L, Li X, Liu F, Sun X, Li C, Zhu Z, Fu Y, Cai H, Wang X, Xie D, Sun C. Control of a key transition from prostrate to erect growth in rice domestication. Nat Genet, 2008, 40: 1360-1364.
doi: 10.1038/ng.197 pmid: 18820699
[11] Yu X L, Ruan M B, Wang B, Yang Y L, Wang S C, Peng M. A homeodomain-leucine zipper I transcription factor, MeHDZ14, regulates internode elongation and leaf rolling in cassava (Manihot esculenta Crantz). Crop J, 2023, 11: 1419-1430.
[12] 雷宁. 木薯TCP转录因子家族的鉴定及MeTCP4的抗逆功能研究. 海南大学硕士学位论文,海南海口, 2018.
Lei N. Identification of Cassava TCP Transcription Factor Family and Study on the Anti Stress Function of MeTCP4. MS Thesis of Hainan University, Haikou, Hainan, China, 2018. (in Chinese with English abstract)
[13] 耿沙, 张建禹, 王晓彤, 任思杨, 毋志浩, 姚远, 李瑞梅, 郭建春, 刘姣, 罗丽娟. 基于CRISPR/Cas9技术创制木薯MeSTP7和MeSTP15双基因突变体. 热带作物学报, 2022, 43: 463-472.
doi: 10.3969/j.issn.1000-2561.2022.03.004
Geng S, Zhang J Y, Wang X D, Ren S Y, Wu Z H, Yao Y, Li R M, Guo J C, Liu J, Luo L H. Creation of cassava MeSTP7 and MeSTP15 dual gene mutants based on CRISPR/Cas9 technology. Chin J Trop Crops, 2022, 43: 463-472 (in Chinese with English abstract).
[14] 徐崟海, 刘佳. IGT基因家族调控作物株型研究进展. 生物技术进展, 2022, 12: 673-682.
doi: 10.19586/j.2095-2341.2022.0103
Xu Y H, Liu J. Research progress in IGT gene family regulation of crop plant type. Curr Biotechnol, 2022, 12: 673-682. (in Chinese with English abstract)
[15] 尚小文, 秦昊, 段玉. 茶树(Camellia sinensis)分枝相关基因家族IGT的鉴定与表达分析. 分子植物育种, 2022, https://kns.cnki.net/kcms/detail/46.1068.S.20220412.1805.028.html.
Shang X W, Qin H, Duan Y. Identification and expression analysis of the IGT family of branching related genes in Camellia sinensis. Mol Plant Breed, 2022, https://kns.cnki.net/kcms/detail/46.1068.S.20220412.1805.028.html (in Chinese with English abstract).
[16] Yoshihara T, Spalding E P. LAZY genes mediate the effects of gravity on auxin gradients and plant architecture. Plant Physiol, 2017, 175: 959-969.
doi: 10.1104/pp.17.00942 pmid: 28821594
[17] Yan H, Yong F S, Xiao B Z. Identification of a gravitropism-deficient mutant in rice. Rice Sci, 2017, 24: 109-118.
doi: 10.1016/j.rsci.2016.06.009
[18] Li P, Wang Y, Qian Q. LAZY1 controls rice shoot gravitropism through regulating polar auxin transport. Cell Res, 2007, 17: 402-410.
doi: 10.1038/cr.2007.38 pmid: 17468779
[19] Dong Z B, Jiang C, Chen X Y. Maize LAZY1 mediates shoot gravitropism and inflorescence development through regulating auxin transport, auxin signaling, and light response. Plant Physiol, 2013, 163: 1306-1322.
doi: 10.1104/pp.113.227314 pmid: 24089437
[20] Yu G C, David S, Zhu H C, Guan Y. Ggtree: an R package for visualization and annotation of phylogenetic trees with their covariates and other associated data. Meth Ecol Evol, 2017, 8: 28-36.
[21] Wilson M C, Mutka A M. Gene expression atlas for the food security crop cassava. New Phytol, 2017, 213: 1632-1641.
doi: 10.1111/nph.14443 pmid: 28116755
[22] 魏胜华, 孟娜. 改良CTAB法提取大戟属药用植物叶片总DNA试验. 湖北农业科学, 2011, 50: 3418-3420.
Wei S H, Meng N. Improved CTAB method for extracting total DNA from leaves of euphorbia medicinal plants. Hubei Agric Sci, 2011, 50: 3418-3420. (in Chinese with English abstract)
[23] Utsumi Y, Utsumi C, Tanaka M. Agrobacterium-mediated cassava transformation for the asian elite variety KU50. Plant Mol Biol, 2021, 109: 271-282.
doi: 10.1007/s11103-021-01212-1 pmid: 34825349
[24] Yoshihara T, Moritoshi I. AtLAZY1 is a signaling component required for gravitropism of the Arabidopsis thaliana inflorescence. Plant J, 2013, 74: 267-279.
[25] Zhang H, Li X, Sang D. PROG1 acts upstream of LAZY1 to regulate rice tiller angle as a repressor. Crop J, 2023, 11: 386-393.
doi: 10.1016/j.cj.2022.11.008
[26] Xia X B, Mi X Z, Jin L, Guo R. CsLAZY1 mediates shoot gravitropism and branch angle in tea plants (Camellia sinensis). BMC Plant Biol, 2021, 21: 243.
doi: 10.1186/s12870-021-03044-z pmid: 34049485
[27] 黄小龙, 孙贵连, 马丹丹. 水稻幼苗酵母单杂文库构建及LAZY1上游调控因子筛选. 生物技术通报, 2023, 39(9): 126-135.
doi: 10.13560/j.cnki.biotech.bull.1985.2023-0001
Huang X L, Sun G L, Ma D D. Construction of rice seedling yeast monohybrid library and screening of LAZY1 upstream regulatory factors. Biotechnol Bull, 2023, 39(9): 126-135. (in Chinese with English abstract)
[28] Yoshihara T, Iino M. Identification of the gravitropism-related rice gene LAZY1 and elucidation of LAZY1-dependent and- independent gravity signaling pathways. Plant Cell Physiol, 2007, 48: 678-688.
doi: 10.1093/pcp/pcm042 pmid: 17412736
[29] Xu D, Qi X, Li J. PzTAC and PzLAZY from a narrow-crown poplar contribute to regulation of branch angles. Plant Physiol Biochem, 2017, 118: 571-578.
[30] Van Overbeek J. Growth substance curvatures of avena in light and dark. J Gene Physiol, 1936, 20: 283-309.
[31] Li P, Wang Y, Qian Q. LAZY1 controls rice shoot gravitropism through regulating polar auxin transport. Cell Res, 2007, 17: 402-410.
doi: 10.1038/cr.2007.38 pmid: 17468779
[32] Li Z, Liang Y, Yuan Y D, Wang L. OsBRXL4 regulates shoot gravitropism and rice tiller angle through affecting LAZY1 nuclear localization. Mol Plant, 2019, 12: 1143-1156.
doi: S1674-2052(19)30200-X pmid: 31200078
[1] SHANG-GUAN Xiao-Xia, YANG Qin-Li, LI Huan-Li. Analysis of mutants developed by CRISPR/Cas9-based GhbHLH71 gene editing in cotton [J]. Acta Agronomica Sinica, 2024, 50(1): 138-148.
[2] YU Xue-Ting, LI Ke, LI Meng-Tao, BAO Ru-Xue, CHEN Xin, WANG Wen-Quan. Interaction identification between protein kinase MeSnRK2.12 and transcription factor MebHLH1 and its relative expression level in cassava [J]. Acta Agronomica Sinica, 2023, 49(9): 2594-2600.
[3] WEI Xin-Yu, ZENG Yue-Hui, YANG Wang-Xing, XIAO Chang-Chun, HOU Xin-Po, HUANG Jian-Hong, ZOU Wen-Guang, XU Xu-Ming. Development of high-quality fragrant indica CMS line by editing Badh2 gene using CRISPR-Cas9 technology in rice (Oryza sativa L.) [J]. Acta Agronomica Sinica, 2023, 49(8): 2144-2159.
[4] XU Zi-Yin, YU Xiao-Ling, ZOU Liang-Ping, ZHAO Ping-Juan, LI Wen-Bin, GENG Meng-Ting, RUAN Meng-Bin. Expression pattern analysis and interaction protein screening of cassava MYB transcription factor MeMYB60 [J]. Acta Agronomica Sinica, 2023, 49(4): 955-965.
[5] LEI Jian-Feng, LI Yue, DAI Pei-Hong, ZHAO Yi, YOU Yang-Zi, JIA Jian-Guo, ZHAO Shuai, QU Yan-Ying, LIU Xiao-Dong. Study on VIGE system mediated by different plant viruses in cotton [J]. Acta Agronomica Sinica, 2023, 49(4): 978-987.
[6] LI Zhao-Wei, MO Zu-Yi, SUN Cong-Ying, SHI Yu, SHANG Ping, LIN Wei-Wei, FAN Kai, LIN Wen-Xiong. Construction of rice mutants by gene editing of OsNAC2d and their response to drought stress [J]. Acta Agronomica Sinica, 2023, 49(2): 365-376.
[7] NIU Zhi-Yuan, QIN Chao, LIU Jun, WANG Hai-Ze, LI Hong-Yu. Function analysis of different Cas9 promoters on the efficiency of CRISPR/ Cas9 system in soybean [J]. Acta Agronomica Sinica, 2023, 49(12): 3227-3238.
[8] CHEN Hui-Xian, LIANG Zhen-Hua, HUANG Zhen-Ling, WEI Wan-Ling, ZHANG Xiu-Fen, YANG Hai-Xia, LI Heng-Rui, HE Wen, LI Tian-Yuan, LAN Xiu, RUAN Li-Xia, CAI Zhao-Qin, NONG Jun-Xin. Transcriptomic profile of key stages of sex differentiation in cassava flowers and discovery of candidate genes related to female flower differentiation [J]. Acta Agronomica Sinica, 2023, 49(12): 3250-3260.
[9] LI Xiang-Chen, SHEN Xu, ZHOU Xin-Cheng, CHEN Xin, WANG Hai-Yan, WANG Wen-Quan. Identification and relative expression levels of PEPC gene family members in cassava [J]. Acta Agronomica Sinica, 2022, 48(12): 3108-3119.
[10] SUN Qian, ZOU Mei-Ling, ZHANG Chen-Ji, JIANG Si-Rong, Eder Jorge de Oliveira, ZHANG Sheng-Kui, XIA Zhi-Qiang, WANG Wen-Quan, LI You-Zhi. Genetic diversity and population structure analysis by SNP and InDel markers of cassava in Brazil [J]. Acta Agronomica Sinica, 2021, 47(1): 42-49.
[11] ZHANG Xiao-Qiong, WANG Xiao-Wen, TIAN Wei-Jiang, ZHANG Xiao-Bo, Sun Ying, LI Yang-Yang, Xie Jia, HE Guang-Hua,SANG Xian-Chun. LAZY1 Regulates the Development of Rice Leaf Angle through BR Pathway [J]. Acta Agron Sin, 2017, 43(12): 1767-1773.
[12] DENG Chang-Zhe,YAO Hui,AN Fei-Fei,LI Kai-Mian,CHEN Song-Bi. Chromoplast Isolation and Its Proteomic Analysis from Cassava Storage Roots [J]. Acta Agron Sin, 2017, 43(09): 1290-1299.
[13] YU Xiao-Ling,RUAN Meng-Bin,WANG Bin,YANG Yi-Ling,WANG Shu-Chang*,PENG Ming*. Cloning and Analysis of Structure and Expression of MeHDZ14 Gene in Cassava [J]. Acta Agron Sin, 2017, 43(08): 1181-1189.
[14] DU Wei-Li,GAO Jie,HU Fu-Liang,GUO De-Lin,ZHANG Gai-Sheng,ZHANG Ren-He*,XUE Ji-Quan. Responses of Drought Stress on Photosynthetic Trait and Osmotic Adjustment in Two Maize Cultivars [J]. Acta Agron Sin, 2013, 39(03): 530-536.
[15] HUANG Qiao-Yi,TANG Shuan-Hu,CHEN Jian-Sheng,ZHANG Fa-Bao,XIE Kai-Zhi,HUANG Xu,JIANG Rui-Ping,LI Ping. Characteristics of Dry Matter Accumulation and Effect of Fertilizer Application in Cassava [J]. Acta Agron Sin, 2013, 39(01): 126-132.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
[1] Li Shaoqing, Li Yangsheng, Wu Fushun, Liao Jianglin, Li Damo. Optimum Fertilization and Its Corresponding Mechanism under Complete Submergence at Booting Stage in Rice[J]. Acta Agronomica Sinica, 2002, 28(01): 115 -120 .
[2] YANG Jian-Chang;ZHANG Jian-Hua;WANG Zhi-Qin;ZH0U Qing-Sen. Changes in Contents of Polyamines in the Flag Leaf and Their Relationship with Drought-resistance of Rice Cultivars under Water Deficiency Stress[J]. Acta Agron Sin, 2004, 30(11): 1069 -1075 .
[3] Yan Mei;Yang Guangsheng;Fu Tingdong;Yan Hongyan. Studies on the Ecotypical Male Sterile-fertile Line of Brassica napus L.Ⅲ. Sensitivity to Temperature of 8-8112AB and Its Inheritance[J]. Acta Agron Sin, 2003, 29(03): 330 -335 .
[4] Wang Yongsheng;Wang Jing;Duan Jingya;Wang Jinfa;Liu Liangshi. Isolation and Genetic Research of a Dwarf Tiilering Mutant Rice[J]. Acta Agron Sin, 2002, 28(02): 235 -239 .
[5] WANG Li-Yan;ZHAO Ke-Fu. Some Physiological Response of Zea mays under Salt-stress[J]. Acta Agron Sin, 2005, 31(02): 264 -268 .
[6] TIAN Meng-Liang;HUNAG Yu-Bi;TAN Gong-Xie;LIU Yong-Jian;RONG Ting-Zhao. Sequence Polymorphism of waxy Genes in Landraces of Waxy Maize from Southwest China[J]. Acta Agron Sin, 2008, 34(05): 729 -736 .
[7] HU Xi-Yuan;LI Jian-Ping;SONG Xi-Fang. Efficiency of Spatial Statistical Analysis in Superior Genotype Selection of Plant Breeding[J]. Acta Agron Sin, 2008, 34(03): 412 -417 .
[8] WANG Yan;QIU Li-Ming;XIE Wen-Juan;HUANG Wei;YE Feng;ZHANG Fu-Chun;MA Ji. Cold Tolerance of Transgenic Tobacco Carrying Gene Encoding Insect Antifreeze Protein[J]. Acta Agron Sin, 2008, 34(03): 397 -402 .
[9] ZHENG Xi;WU Jian-Guo;LOU Xiang-Yang;XU Hai-Ming;SHI Chun-Hai. Mapping and Analysis of QTLs on Maternal and Endosperm Genomes for Histidine and Arginine in Rice (Oryza sativa L.) across Environments[J]. Acta Agron Sin, 2008, 34(03): 369 -375 .
[10] XING Guang-Nan, ZHOU Bin, ZHAO Tuan-Jie, YU De-Yue, XING Han, HEN Shou-Yi, GAI Jun-Yi. Mapping QTLs of Resistance to Megacota cribraria (Fabricius) in Soybean[J]. Acta Agronomica Sinica, 2008, 34(03): 361 -368 .