水稻OsLHT1在拟南芥中的表达对植物生长的效应初探
Effect of OsLHT1 Expression on Plant Growth in Arabidopsis thaliana

DOI: 10.12677/BR.2021.102022, PP. 146-154

Keywords: 拟南芥，水稻，OsLHT1，异源表达
Arabidopsis thaliana, Rice, OsLHT1, Heterologous Expression

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Abstract:

水稻氨基酸转运蛋白OsLHT1与拟南芥AtLHT1的蛋白质序列具有较高的同源性，经比对发现相似度达78%。拟南芥氨基酸转运蛋白AtLHT1突变体(atlht1)相比较野生型生长矮小，在开花后期出现分蘖数和果荚数少、早衰等症状。为了探究水稻OsLHT1在植物生长发育中的分子生物学效应/功能，本研究通过RT-PCR的方法，在水稻中克隆获得了OsLHT1全长ORF片段，构建了35s-OsLHT1超表达载体，异源表达于拟南芥atlht1突变体中，并对OsLHT1在水稻重要的生育时期做了基因表达分析。结果表明，转基因植株(atlht1 + 35s-OsLHT1)能够恢复拟南芥atlht1的生长表型，分枝数与果荚数均比突变体增多，转录水平分析结果表明OsLHT1在水稻重要生育时期都有表达，且在分蘖期的根部和老叶的表达量高，推测OsLHT1在植物生长发育过程中起到了关键作用。
Rice amino acid transporter OsLHT1 is highly homologous to Arabidopsis AtLHT1, with sharing a similarity of 78% at the protein level. The Arabidopsis amino acid transporter AtLHT1 mutant (atlht1), compared with the wild-type, shows a growth in small size, less upper part branches and siliques, as well as early senescence at the late flowering stage. In order to explore the molecular biological effects of OsLHT1 on plant growth and development，we cloned the full length open reading frame (ORF) of OsLHT1 using RT-PCR, constructed a 35s-OsLHT1 overexpression vector, heterologously expressed OsLHT1 in the Arabidopsis atlht1 mutant, and analyzed the expression of OsLHT1 in rice at significant growth stages. The result showed that transgenic plants (atlht1 + 35s-OsLHT1) indeed displayed a restored growth phenotype related to atlht1 mutant, the number of the upper branches and pods was higher than that of mutants; moreover, gene expression study showed that OsLHT1 was expressed at almost all important growth stages of rice, but the highest expression level was observed in roots of the tillering stage and old leaves, thus proposing the critical role for OsLHT1 in plant growth and development.

References

[1]	Coruzzi, G. and Bush, D.R. (2001) Nitrogen and Carbon Nutrient and Metabolite Signaling in Plants. Plant Physiology, 125, 61-64. https://doi.org/10.1104/pp.125.1.61
[2]	Jones, D.L., Owen, A.G. and Farrar, J.F. (2002) Simple Method to Enable the High Resolution Determination of Total Free Amino Acids in Soil Solutions and Soil Extracts. Soil Biology & Biochemistry, 34, 1893-1902. https://doi.org/10.1016/S0038-0717(02)00203-1
[3]	Svennerstam, H., J？mtg？rd, S., Ahmad, I., et al. (2011) Transporters in Arabidopsis Roots Mediating Uptake of Amino Acids at Naturally Occurring Concentrations. New Phytologist, 191, 459-467. https://doi.org/10.1111/j.1469-8137.2011.03699.x
[4]	Watanabe, M., Balazadeh, S., Tohge, T., et al. (2013) Comprehensive Dissection of Spatiotemporal Metabolic Shifts in Primary, Secondary, and Lipid Metabolism during Developmental Senescence in Arabidopsis. Plant Physiology, 162, 1290-1310. https://doi.org/10.1104/pp.113.217380
[5]	Hildebrandt, T.M., Nesi, A.N., Araújo, W.L. and Braun, H.-P. (2015) Amino Acid Catabolism in Plants. Molecular Plant, 8, 1563-1579. https://doi.org/10.1016/j.molp.2015.09.005
[6]	Walch-Liu, P., Liu, L., Remans, T., et al. (2006) Evidence That L-Glutamate Can Act as an Exogenous Signal to Modulate Root Growth and Branching in Arabidopsis thaliana. Plant and Cell Physiology, 47, 1045-1057. https://doi.org/10.1093/pcp/pcj075
[7]	Li, Z.C. and Bush, D.R. (1991) DeltapH-Dependent Amino Acid Transport into Plasma Membrane Vesicles Isolated from Sugar Beet (Beta vulgaris L.) Leaves: II. Evidence for Multiple Aliphatic, Neutral Amino Acid Symports. Plant Physiology, 96, 1338-1344. https://doi.org/10.1104/pp.96.4.1338
[8]	Weston, K., Hall, J.L. and Williams, L.E. (1995) Characterization of Amino-Acid Transport in Ricinus communis Roots Using Isolated Membrane Vesicles. Planta, 196, 166-173. https://doi.org/10.1007/BF00193230
[9]	Wyse, R.E. and Komor, E. (1984) Mechanism of Amino Acid Uptake by Sugarcane Suspension Cells. Plant Physiology, 76, 865-870. https://doi.org/10.1104/pp.76.4.865
[10]	Frommer, W.B., Hummel, S. and Riesmeier, J.W. (1993) Expression Cloning in Yeast of a cDNA Encoding a Broad Specificity Amino Acid Permease from Arabidopsis thaliana. Pro-ceedings of the National Academy of Sciences, 90, 5944-5948. https://doi.org/10.1073/pnas.90.13.5944
[11]	Hsu, L.C., Chiou, T.J., Chen, L, and Bush, D.R. (1993) Cloning a Plant Amino Acid Transporter by Functional Complementation of a Yeast Amino Acid Transport Mutant. Proceedings of the National Academy of Sciences, 90, 7441-7445. https://doi.org/10.1073/pnas.90.16.7441
[12]	Pratelli, R. and Pilot, G. (2014) Regulation of Amino Acid Metabolic Enzymes and Transporters in Plants. Journal of Experimental Botany, 65, 5535-5556. https://doi.org/10.1093/jxb/eru320
[13]	Tegeder, M. (2012) Transporters for Amino Acids in Plant Cells: Some Functions and Many Unknowns. Current Opinion in Plant Biology, 15, 315-321. https://doi.org/10.1016/j.pbi.2012.02.001
[14]	Schwacke, R., Schneider, A., van der Graaff, E., et al. (2003) ARAMEMNON, a Novel Database for Arabidopsis Integral Membrane Proteins. Plant Physiology, 131, 16-26. https://doi.org/10.1104/pp.011577
[15]	Rentsch, D., Schmidt, S. and Tegeder, M. (2007) Transporters for Uptake and Allocation of Organic Nitrogen Compounds in Plants. FEBS Letters, 581, 2281-2289. https://doi.org/10.1016/j.febslet.2007.04.013
[16]	Chen, L. and Bush, D.R. (1997) LHT1, A Lysine- and Histidine-Specific Amino Acid Transporter in Arabidopsis. Plant Physiology, 115, 1127-1134. https://doi.org/10.1104/pp.115.3.1127
[17]	Ganeteg, U., Ahmad, I., J？mtg？rd, S., et al. (2016) Amino Acid Transporter Mutants of Arabidopsis Provides Evidence That a Non-Mycorrhizal Plant Acquires Organic Nitrogen from Agricultural Soil. Plant, Cell & Environment, 40, 413-423. https://doi.org/10.1111/pce.12881
[18]	Hirner, A., Ladwig, F., Stransky, H., et al. (2006) Arabidopsis LHT1 Is a High-Affinity Transporter for Cellular Amino Acid Uptake in Both Root Epidermis and Leaf Mesophyll. The Plant Cell, 18, 1931-1946. https://doi.org/10.1105/tpc.106.041012
[19]	Perchlik, M., Foster, J. and Tegeder, M. (2014) Different and Overlapping Functions of Arabidopsis LHT6 and AAP1 Transporters in Root Amino Acid Uptake. Journal of Experimental Botany, 65, 5193-5204. https://doi.org/10.1093/jxb/eru278
[20]	Wang, X., Yang, G., Shi, M., et al. (2019) Disruption of an Amino Acid Transporter LHT1 Leads to Growth Inhibition and Low Yields in Rice. BMC Plant Biology, 19, Article No. 268. https://doi.org/10.1186/s12870-019-1885-9
[21]	Guo, N., Hu, J., Yan, M., et al. (2020) Oryza sativa Ly-sine-Histidine-Type Transporter 1 Functions in Root Uptake and Root-to-Shoot Allocation of Amino Acids in Rice. The Plant Journal, 103, 395-411. https://doi.org/10.1111/tpj.14742
[22]	Guo, N., Gu, M., Hu, J., Qu, H. and Xu, G. (2020) Rice OsLHT1 Functions in Leaf-to-Panicle Nitrogen Allocation for Grain Yield and Quality. Frontiers in Plant Science, 11, 1150. https://doi.org/10.3389/fpls.2020.01150
[23]	Wang, W.H., K？hler, B., Cao, F.Q., et al. (2011) Rice DUR3 Mediates High-Affinity Urea Transport and Plays an Effective Role in Improvement of Urea Acquisition and Utilization When Expressed in Arabidopsis. New Phytologist, 193, 432-444. https://doi.org/10.1111/j.1469-8137.2011.03929.x
[24]	Clough, S.J. and Bent, A.F. (1998) Floral Dip: A Simplified Method for Agrobacterium-Mediated Transformation of Arabidopsis thaliana. The Plant Journal, 16, 735-743. https://doi.org/10.1046/j.1365-313x.1998.00343.x

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