Browse > Article

Cloning and Characterization of ${\Delta}^1$-Pyrroline-5-Carboxylate Synthetase Genes and Identification of Point Mutants in Medicago truncatula  

Song, Ki-Hoon (Department of Life Science, Sogang University)
Song, Dae-Hae (Department of Life Science, Sogang University)
Lee, Jeong-Ran (Department of Life Science, Sogang University)
Kim, Goon-Bo (Department of Life Science, Sogang University)
Choi, Hong-Kyu (Department of Plant Pathology, University of California)
Penmetsa, R. Varma (Department of Plant Pathology, University of California)
Nam, Young-Woo (Department of Life Science, Sogang University)
Publication Information
KOREAN JOURNAL OF CROP SCIENCE / v.52, no.4, 2007 , pp. 458-468 More about this Journal
Abstract
To tolerate environmentally adverse conditions such as cold, drought, and salinity, plants often synthesize and accumulate proline in cells as compatible osmolytes. ${\Delta}^1$-Pyrroline-5-carboxylate synthetase(P5CS) catalyzes the rate-limiting step of proline biosynthesis from glutamate. Two complete genes, MtP5CS1 and MtP5CS2, were isolated from the model legume Medicago truncatula by cDNA cloning and bacterial artificial chromosome library screening. Nucleotide sequence analysis showed that both genes consisted of 20 exons and 19 introns. Alignment of the predicted amino acid sequences revealed high similarities with P5CS proteins from other plant species. The two MtP5CS genes were expressed in response to high salt and low temperature treatments. Semi-quantitative reverse transcription-polymerase chain reaction showed that MtP5CS1 was expressed earlier than MtP5CS2, indicating differential regulation of the two genes. To evaluate the reverse genetic effects of nucleotide changes on MtP5CS function, a Targeting Induced Local Lesions in Genomes approach was taken. Three mutants each were isolated for MtP5CS1 and MtP5CS2, of which a P5CS2 nonsense mutant carrying a codon change from arginine to stop was expected to bring translation to premature termination. These provide a valuable genetic resource with which to determine the function of the P5CS genes in environmental stress responses of legume crops.
Keywords
medicago truncatula; model legume; stress response; proline; ${\Delta}^1$-pyrroline-5-carboxylate synthetase; nonsense mutant;
Citations & Related Records
Times Cited By KSCI : 2  (Citation Analysis)
연도 인용수 순위
1 Armengaud, P., L. Thiery, N. Buhot, G. Grenier-De March, and A. Savoure. 2004. Transcriptional regulation of proline biosynthesis in Medicago truncatula reveals developmental and environmental specific features. Physiol. Plant. 120 : 442-450   DOI   ScienceOn
2 Baulcombe, D. 2004. RNA silencing in plants. Nature 431 : 356-363   DOI   ScienceOn
3 Buchannan, B. B., W. Gruissem, and R. L. Jones. 2000. Biochemistry and Molecular Biology of Plants. American Society of Plant Biologists, pp. 406-409
4 Nam, Y.-W., R. V. Penmetsa, G. Endre, P. Uribe, D. Kim, and D. R. Cook. 1999. Construction of a bacterial artificial chromosome library of Medicago truncatula and identification of clones for ethylene responsive genes. Theor. Appl. Genet. 98 : 638-646   DOI
5 Yoshiba, Y., T. Kiyosue, T. Katagiri, H. Ueda, T. Mizoguchi, K. Yamaguchi-Shinozaki, K. Wada, Y. Harada, and K. Shinozaki. 1995. Correlation between the induction of a gene for $\Delta^{1}$-pyrroline-5-carboxylate synthetase and the accumulation of proline in Arabidopsis thaliana under osmotic stress. Plant J. 7 : 751-760   DOI   ScienceOn
6 Park, S.-Y. and Y.-W. Nam. 2006. Construction of bacterial artificial chromosome library containing large BamHI genomic fragments from Medicago truncatula and identification of clones linked to hyper-nodulating genes. J. Microbiol. Biotechnol. 16 : 256-263   과학기술학회마을
7 Zhu, H. Y., D. J. Kim, J. M. Baek, H. K. Choi, L. Ellis, H. Kuester, W. R. McCombie, H. M. Peng, and D. R. Cook. 2003. Syntenic relationships between Medicago truncatula and Arabidopsis thaliana reveal extensive divergence of genome organization. Plant Physiol. 131 : 1018-1026   DOI   ScienceOn
8 Csonka, L. N. and A. D. Hanson. 1991. Prokaryotic osmoregulation: genetics and physiology. Annu. Rev. Microbiol. 45 : 569-606   DOI   ScienceOn
9 Bouche, N. and D. Bouchez. 2001. Arabidopsis gene knockout: phenotypes wanted. Curr. Opin. Plant Biol. 4 : 111-117   DOI   ScienceOn
10 Trieu, A. T., S. H. Burleigh, I. V. Kardailsky, I. E. Maldonado-Mendoza, W. K. Versaw, L. A. Blaylock, H. Shin, T. J. Chiou, H. Katagi, G. R. Dewbre, D. Weigel, and M. J. Harrison. 2000. Transformation of Medicago truncatula via infiltration of seedlings or flowering plants with Agrobacterium. Plant J. 22: 531-541   DOI   ScienceOn
11 Thompson, J. D., D. G. Higgins, and T. J. Gibson. 1994. CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22 : 4673-4680   DOI
12 Till, B. J., C. Burtner, L. Comai, and S. Henikoff. 2004. Mismatch cleavage by single-strand specific nucleases. Nucleic Acids Res. 32 : 2632-2641   DOI   ScienceOn
13 Troll, W. and J. Lindsley. 1955. A photometric method for the determination of proline. J. Biol. Chem. 215 : 655-660
14 Penmetsa, R. V. and D. R. Cook. 1997. A legume ethylene-insensitive mutant hyperinfected by its rhizobial symbiont. Science 275 : 527-530   DOI   ScienceOn
15 Igarashi, Y., Y. Yoshiba, Y. Sanada, K. Yamaguchi-Shinozaki, K. Wada, and K. Shinozaki. 1997. Characterization of the gene for ${\Delta}^1-pyrroline-5-carboxylate$ synthetase and correlation between the expression of the gene and salt tolerance in Oryza sativa L. Plant Mol. Biol. 33 : 857-865   DOI   ScienceOn
16 Strizhov, N., E. Abraham, L. Okresz, S. Blickling, A. Zilberstein, J. Schell, C. Koncz, and L. Szabados. 1997. Differential expression of two P5CS genes controlling proline accumulation during salt-stress requires ABA and is regulated by ABA1, ABI1 and AXR2 in Arabidopsis. Plant J. 12 : 557-569   DOI   ScienceOn
17 Blondon, F., D. Marie, S. Brown, and A. Kondorosi. 1994. Genome size and base composition in Medicago sativa and Medicago truncatula species. Genome 37 : 264-270   DOI
18 Barker, D. G., S. Bianchi, F. Blondon, Y. Dattee, G. Duc, S. Essad, P. Flament, P. Gallusci, G. Genier, P. Guy, X. Muel, J. Tourneur, J. Denarie, and T. Huguet. 1990. Medicago truncatula, a model plant for studying the molecular genetics of the Rhizobium-legume symbiosis. Plant Mol. Biol. Rep. 8 : 40-49
19 Deuschle, K., D. Funck, H. Hellmann, K. Daschner, S. Binder, W. B. Frommer. 2001. A nuclear gene encoding mitochondrial delta-pyrroline-5-carboxylate dehydrogenase and its potential role in protection from proline toxicity. Plant J. 27 : 345-356   DOI   ScienceOn
20 VandenBosch, K. A. and G. Stacey. 2003. Summaries of legume genomics projects from around the globe. Community resources for crops and models. Plant Physiol. 131 : 840-865   DOI   ScienceOn
21 Kiyosue, T., Y. Yoshiba, K. Yamaguchi-Slnnozaki, and K. Shinozaki. 1996. A nuclear gene encoding mitochondrial proline dehydrogenase, an enzyme involved in proline metabolism, is upregulated by proline but downregulated by dehydration in Arabidopsis. Plant Cell 8 : 1323-1335   DOI   ScienceOn
22 McCallum, C. M., L. Comai, E. A. Greene, and S. Henikoff. 2000. Targeting induced local lesions IN genomes (TILLING) for plant functional genomics. Plant Physiol. 123 : 439-442   DOI   ScienceOn
23 Perry, J. A., T. L. Wang, T. J. Welham, S. Gardner, J. M. Pike, S. Yoshida, and M. Parniske. 2003. A TILLING reverse genetics tool and a web-accessible collection of mutants of the legume Lotus japonicus. Plant Physiol. 131 : 866-871   DOI   ScienceOn
24 Neff, M. M., J. D. Neff, J. Chory, and A. E. Pepper. 1998. dCAPS, a simple technique for the genetic analysis of single nucleotide polymorphisms: experimental applications in Arabidopsis thaliana genetics. Plant J. 14 : 387-392   DOI   ScienceOn
25 Ginzberg, I., H. Stein, Y. Kapulnik, L. Szabados, N. Strizhov, J. Schell, C. Koncz, and A. Zilberstein. 1998. Isolation and characterization of two different cDNAs of $\Delta^{1}$-pyrroline-5-carboxylate synthase in alfalfa transcriptionally induced upon salt stress. Plant Mol. Biol. 38 : 755-764   DOI   ScienceOn
26 Comai, L. and S. Henikoff. 2006. TILLING: practical single-nucleotide mutation discovery. Plant J. 45 : 684-694   DOI   ScienceOn
27 Seki, M., A. Kamei, K. Yamaguchi-Shinozaki, and K. Shinozaki. 2003. Molecular responses to drought, salinity and frost: common and different paths for plant protection. Curr. Opin. Biotechnol. 14 : 194-199   DOI   ScienceOn
28 Choi, H. K., D. Kim, T. Uhm, E. Limpens, H. Lim, P. Kalo, R. V. Penmetsa, A. Seres, O. Kulikova, T. Bisseling, G. B. Kiss, and D. R. Cook. 2004. A sequence-based genetic map of Medicago truncatula and comparison of marker co-linearity with Medicago sativa. Genetics 166 : 1463-1502   DOI
29 Till, B. J., T. Zerr, L. Comai, and S. Henikoff. 2006. A protocol for TILLING and Ecotilling in plants and animals. Nat. Protocols 1 : 2465-2477   DOI   ScienceOn
30 Song, K.-H. and Y.-W. Nam. 2005. Genomic organization and differential expression of two polygalacturonase-inhibiting protein genes from Medicago truncatula. J. Plant Biol. 48 : 467-478   과학기술학회마을   DOI