DOI QR코드

DOI QR Code

Molecular cloning and expression analysis of the first two key genes through 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway from Pyropia haitanensis (Bangiales, Rhodophyta)

  • Du, Yu (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • Guan, Jian (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • Xu, Ruijun (School for Radiological & Interdisciplinary Sciences, Soochow University) ;
  • Liu, Xin (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • Shen, Weijie (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • Ma, Yafeng (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • He, Yuan (Department of Cell Biology, School of Biology and Basic Medical, Soochow University) ;
  • Shen, Songdong (Department of Cell Biology, School of Biology and Basic Medical, Soochow University)
  • 투고 : 2017.08.27
  • 심사 : 2017.10.02
  • 발행 : 2017.12.15

초록

Pyropia haitanensis (T. J. Chang et B. F. Zheng) N. Kikuchi et M. Miyata is one of the most commercially useful macroalgae cultivated in southeastern China. In red algae, the biosynthesis of terpenoids through 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway can produce a direct influence on the synthesis of many biologically important metabolites. In this study, two genes of cDNAs, 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and 1-deoxy-D-xylulose-5-phosphate reductase (DXR), which encoding the first two rate-limiting enzymes among MEP pathway were cloned from P. haitanensis. The cDNAs of P. haitanensis DXS (PhDXS) and DXR (PhDXR) both contained complete open reading frames encoding polypeptides of 764 and 426 amino acids residues, separately. The expression analysis showed that PhDXS was significant differently expressed between leafy thallus and conchocelis as PhDXR been non-significant. Additionally, expression of PhDXR and its downstream gene geranylgeranyl diphosphate synthase were both inhibited by fosmidomycin significantly. Meanwhile, we constructed types of phylogenetic trees through different algae and higher plants DXS and DXR encoding amino acid sequences, as a result we found tree clustering consequences basically in line with the "Cavalier-Smith endosymbiotic theory." Whereupon, we speculated that in red algae, there existed only complete MEP pathway to meet needs of terpenoids synthesis for themselves; Terpenoids synthesis of red algae derivatives through mevalonate pathway came from two or more times endosymbiosis of heterotrophic eukaryotic parasitifer. This study demonstrated that PhDXS and PhDXR could play significant roles in terpenoids biosynthesis at molecular levels. Meanwhile, as nuclear genes among MEP pathway, PhDXS and PhDXR could provide a new way of thinking to research the problem of chromalveolata biological evolution.

키워드

참고문헌

  1. Adl, S. M., Simpson, A. G., Farmer, M. A., Andersen, R. A., Anderson, O. R., Barta, J. R., Bowser, S. S., Brugerolle, G., Fensome, R. A., Fredericq, S., James, T. Y., Karpov, S., Kugrens, P., Krug, J., Lane, C. E., Lewis, L. A., Lodge, J., Lynn, D. H., Mann, D. G., McCourt, R. M., Mendoza, L., Moestrup, O., Mozley-Standridge, S. E., Nerad, T. A., Shearer, C. A., Smirnov, A. V., Spiegel, F. W. & Taylor, M. F. 2005. The new higher level classification of eukaryotes with emphasis on the taxonomy of protists. J. Eukaryot. Microbiol. 52:399-451. https://doi.org/10.1111/j.1550-7408.2005.00053.x
  2. Bennett, A. & Bogorad, L. 1973. Complementary chromatic adaptation in a filamentous blue-green alga. J. Cell. Biol. 58:1245-1257.
  3. Blouin, N. A., Brodie, J. A., Grossman, A. C., Xu, P. & Brawley, S. H. 2011. Porphyra: a marine crop shaped by stress. Trends. Plant. Sci. 16:29-37. https://doi.org/10.1016/j.tplants.2010.10.004
  4. Bowler, C., Allen, A. E., Badger, J. H., Grimwood, J., Jabbari, K., Kuo, A., Maheswari, U., Martens, C., Maumus, F. & Otillar, R. P. 2008. The phaeodactylum genome reveals the evolutionary history of diatom genomes. Nature 456:239-244. https://doi.org/10.1038/nature07410
  5. Carretero-Paulet, L., Cairo, A., Botella-Pavia, P., Besumbes, O., Campos, N., Boronat, A. & Rodriguez-Concepcion, M. 2006. Enhanced flux through the methylerythritol 4-phosphate pathway in Arabidopsis plants overexpressing deoxyxylulose 5-phosphate reductoisomerase. Plant Mol. Biol. 62:683-695. https://doi.org/10.1007/s11103-006-9051-9
  6. Carretero-Paulet, L., Cairo, A., Talavera, D., Saura, A., Imperial, S., Rodriguez-Concepcion, M., Campos, N. & Boronat, A. 2013. Functional and evolutionary analysis of DXL1, a non-essential gene encoding a 1-deoxy-D-xylulose 5-phosphate synthase like protein in Arabidopsis thaliana. Gene 524:40-53. https://doi.org/10.1016/j.gene.2012.10.071
  7. Cavalier-Smith, T. 1999. Principles of protein and lipid targeting in secondary symbiogenesis: euglenoid, dinoflagellate, and sporozoan plastid origins and the eukaryote family tree. J. Eukaryot. Microbiol. 46:347-366. https://doi.org/10.1111/j.1550-7408.1999.tb04614.x
  8. Chan, C. X., Blouin, N. A., Zhuang, Y., Zäuner, S., Prochnik, S. E., Lindquist, E., Lin, S., Benning, C., Lohr, M., Yarish, C., Gantt, E., Grossman, A. R., Lu, S., Müller, K., Stiller, J. W., Brawley, S. H. & Bhattacharya, D. 2012. Porphyra (Bangiophyceae) transcriptomes provide insights into red algal development and metabolism. J. Phycol. 48:1328-1342. https://doi.org/10.1111/j.1529-8817.2012.01229.x
  9. Chen, C., Dai, Z., Xu, Y., Ji, D. & Xie, C. 2016. Cloning, expression, and characterization of carbonic anhydrase genes from Pyropia haitanensis (Bangiales, Rhodophyta). J. Appl. Phycol. 28:1403-1417. https://doi.org/10.1007/s10811-015-0646-x
  10. Cock, J. M., Sterck, L., Rouze, P., Scornet, D., Allen, A. E., Amoutzias, G., Anthouard, V., Artiguenave, F., Aury, J. M. & Badger, J. H. 2010. The Ectocarpus genome and the independent evolution of multicellularity in brown algae. Nature 465:617-621. https://doi.org/10.1038/nature09016
  11. Cordoba, E., Porta, H., Arroyo, A., San Roman, C., Medina, L., Rodriguez-Concepcion, M. & Leon, P. 2011. Functional characterization of the three genes encoding 1-deoxy-D-xylulose 5-phosphate synthase in maize. J. Exp. Bot. 62:2023-2038. https://doi.org/10.1093/jxb/erq393
  12. Davies, F. K., Jinkerson, R. E. & Posewitz, M. C. 2015. Toward a photosynthetic microbial platform for terpenoid engineering. Photosynth. Res. 123:265-284. https://doi.org/10.1007/s11120-014-9979-6
  13. de Oliveira, L. S., Gregoracci, G. B., Silva, G. G. Z., Salgado, L. T., Filho, G. A., Alves-Ferreira, M., Pereira, R. C. & Thompson, F. L. 2012. Transcriptomic analysis of the red seaweed Laurencia dendroidea (Florideophyceae, Rhodophyta) and its microbiome. BMC Genomics 13:487. https://doi.org/10.1186/1471-2164-13-487
  14. Frommolt, R., Werner, S., Paulsen, H., Goss, R., Wilhelm, C., Zauner, S., Maier, U. G., Grossman, A. R., Bhattacharya, D. & Lohr, M. 2008. Ancient recruitment by chromists of green algal genes encoding enzymes for carotenoid biosynthesis. Mol. Biol. Evol. 25:2653-2667. https://doi.org/10.1093/molbev/msn206
  15. Grauvogel, C. & Petersen, J. 2007. Isoprenoid biosynthesis authenticates the classification of the green alga Mesostigma viride as an ancient streptophyte. Gene 396:125-133. https://doi.org/10.1016/j.gene.2007.02.020
  16. Hans, J., Hause, B., Strack, D. & Walter, M. H. 2004. Cloning, characterization, and immunolocalization of a mycorrhiza-inducible 1-deoxy-d-xylulose 5-phosphate reductoisomerase in arbuscule-containing cells of maize. Plant Physiol. 134:614-624. https://doi.org/10.1104/pp.103.032342
  17. Huang, W., Ye, J., Zhang, J., Lin, Y., He, M. & Huang, J. 2016. Transcriptome analysis of Chlorella zofingiensis to identify genes and their expressions involved in astaxanthin and triacylglycerol biosynthesis. Algal Res. 17:236-243. https://doi.org/10.1016/j.algal.2016.05.015
  18. Kuzuyama, T., Shimizu, T., Takahashi, S. & Seto, H. 1998. Fosmidomycin, a specific inhibitor of 1-deoxy-d-xylulose 5-phosphate reductoisomerase in the nonmevalonate pathway for terpenoid biosynthesis. Tetrahedron. Lett. 39:7913-7916. https://doi.org/10.1016/S0040-4039(98)01755-9
  19. Liu, J., Xu, Y., Liang, L. & Wei, J. 2015. Molecular cloning, characterization and expression analysis of the gene encoding 1-deoxy-D-xylulose 5-phosphate reductoisomerase from Aquilaria sinensis (Lour.) Gilg. J. Genet. 94:239-249. https://doi.org/10.1007/s12041-015-0521-1
  20. Lohr, M., Schwender, J. & Polle, J. E. W. 2012. Isoprenoid biosynthesis in eukaryotic phototrophs: a spotlight on algae. Plant Sci. 185-186:9-22. https://doi.org/10.1016/j.plantsci.2011.07.018
  21. Luo, Q., Zhu, Z., Zhu, Z., Yang, R., Qian, F., Chen, H. & Yan, X. 2014. Different responses to heat shock stress revealed heteromorphic adaptation strategy of Pyropia haitanensis (Bangiales, Rhodophyta). PLoS ONE 9:e94354. https://doi.org/10.1371/journal.pone.0094354
  22. Masse, G., Belt, S. T., Rowland, S. J. & Rohmer, M. 2004. Isoprenoid biosynthesis in the diatoms Rhizosolenia setigera (Brightwell) and Haslea ostrearia (Simonsen). Proc. Natl. Acad. Sci. U. S. A. 101:4413-4418. https://doi.org/10.1073/pnas.0400902101
  23. Matsuzaki, M., Misumi, O., Shin-I, T., Maruyama, S., Takahara, M., Miyagishima, S. Y., Mori, T., Nishida, K., Yagisawa, F., Nishida, K., Yoshida, Y., Nishimura, Y., Nakao, S., Kobayashi, T., Momoyama, Y., Higashiyama, T., Minoda, A., Sano, M., Nomoto, H., Oishi, K., Hayashi, H., Ohta, F., Nishizaka, S., Haga, S., Miura, S., Morishita, T., Kabeya, Y., Terasawa, K., Suzuki, Y., Ishii, Y., Asakawa, S., Takano, H., Ohta, N., Kuroiwa, H., Tanaka, K., Shimizu, N., Sugano, S., Sato, N., Nozaki, H., Ogasawara, N., Kohara, Y. & Kuroiwa, T. 2004. Genome sequence of the ultrasmall unicellular red alga Cyanidioschyzon merolae 10D. Nature 428:653-657. https://doi.org/10.1038/nature02398
  24. Okada, K., Saito, T., Nakagawa, T., Kawamukai, M. & Kamiya, Y. 2000. Five geranylgeranyl diphosphate synthases expressed in different organs are localized into three subcellular compartments in Arabidopsis. Plant Physiol. 122:1045-1056. https://doi.org/10.1104/pp.122.4.1045
  25. Paniagua-Michel, J., Capa-Robles, W., Olmos-Soto, J. & Gutierrez-Millan, L. E. 2009. The carotenogenesis pathway via the isoprenoid-${\beta}$-carotene interference approach in a new strain of Dunaliella salina isolated from Baja California Mexico. Mar. Drugs 7:45-56. https://doi.org/10.3390/md7010045
  26. Pattanaik, B. & Lindberg, P. 2015. Terpenoids and their biosynthesis in cyanobacteria. Life (Basel) 5:269-293.
  27. Peng, G., Wang, C., Song, S., Fu, X., Azam, M., Grierson, D. & Xu, C. 2013. The role of 1-deoxy-d-xylulose-5-phosphate synthase and phytoene synthase gene family in citrus carotenoid accumulation. Plant Physiol. Biochem. 71:67-76. https://doi.org/10.1016/j.plaphy.2013.06.031
  28. Reyes-Prieto, A., Hackett, J. D., Soares, M. B., Bonaldo, M. F. & Bhattacharya, D. 2006. Cyanobacterial contribution to algal nuclear genomes is primarily limited to plastid functions. Curr. Biol. 16:2320-2325. https://doi.org/10.1016/j.cub.2006.09.063
  29. Rodriguez-Concepcion, M., Ahumada, I., Diez-Juez, E., Sauret-Güeto, S., Lois, L. M., Gallego, F., Carretero-Paulet, L., Campos, N. & Boronat, A. 2001. 1-Deoxy-Dxylulose 5-phosphate reductoisomerase and plastid isoprenoid biosynthesis during tomato fruit ripening. Plant J. 27:213-222. https://doi.org/10.1046/j.1365-313x.2001.01089.x
  30. Scolnik, P. A. & Bartley, G. E. 1994. Nucleotide sequence of an Arabidopsis cDNA for geranylgeranyl pyrophosphate synthase. Plant Physiol. 104:1469-1470. https://doi.org/10.1104/pp.104.4.1469
  31. Slamovits, C. H. & Keeling, P. J. 2008. Plastid-derived genes in the nonphotosynthetic alveolate Oxyrrhis marina. Mol. Biol. Evol. 25:1297-1306. https://doi.org/10.1093/molbev/msn075
  32. Sutherland, J. E., Lindstrom, S. C., Nelson, W. A., Brodie, J., Lynch, M. D. J., Hwang, M. S., Choi, H. -G., Miyata, M., Kikuchi, N., Oliveira, M. C., Farr, T., Neefus, C., Mols-Mortensen, A., Milstein, D. & Müller, K. M. 2011. A new look at an ancient order: generic revision of the Bangiales (Rhodophyta). J. Phycol. 47:1131-1151. https://doi.org/10.1111/j.1529-8817.2011.01052.x
  33. Tong, Y., Su, P., Zhao, Y., Zhang, M., Wang, X., Liu, Y., Zhang, X., Gao, W. & Huang, L. 2015. Molecular cloning and characterization of DXS and DXR genes in the terpenoid biosynthetic pathway of Tripterygium wilfordii. Int. J. Mol. Sci. 16:25516-25535. https://doi.org/10.3390/ijms161025516
  34. Vranova, E., Coman, D. & Gruissem, W. 2013. Network analysis of the MVA and MEP pathways for isoprenoid synthesis. Annu. Rev. Plant. Biol. 64:665-700. https://doi.org/10.1146/annurev-arplant-050312-120116
  35. Wang, L., Mao, Y., Kong, F., Cao, M. & Sun, P. 2015. Genomewide expression profiles of Pyropia haitanensis in response to osmotic stress by using deep sequencing technology. BMC Genomics 16:1012. https://doi.org/10.1186/s12864-015-2226-5
  36. Wellburn, A. R. 1994. The spectral determination of chlorophylls a, and b, as well as total carotenoids, using various solvents with spectrophotometers of different resolution. J. Plant. Physiol. 144:307-313. https://doi.org/10.1016/S0176-1617(11)81192-2
  37. Xiang, S., Usunow, G., Lange, G., Busch, M. & Tong, L. 2007. Crystal structure of 1-deoxy-d-xylulose 5-phosphate synthase, a crucial enzyme for isoprenoids biosynthesis. J. Biol. Chem. 282:2676-2682. https://doi.org/10.1074/jbc.M610235200
  38. Xie, C., Li, B., Xu, Y., Ji, D. & Chen, C. 2013. Characterization of the global transcriptome for Pyropia haitanensis (Bangiales, Rhodophyta) and development of cSSR markers. BMC Genomics 14:107. https://doi.org/10.1186/1471-2164-14-107
  39. Xu, D., Qiao, H., Zhu, J., Xu, P., Liang, C., Zhang, X., Ye, N. & Yang, W. 2012. Assessment of photosynthetic performance of Porphyra yezoensis (Bangiales, Rhodophyta) in conchocelis phase. J. Phycol. 48:467-470. https://doi.org/10.1111/j.1529-8817.2012.01121.x
  40. Xu, Y., Liu, J., Liang, L., Yang, X., Zhang, Z., Gao, Z., Sui, C. & Wei, J. 2014. Molecular cloning and characterization of three cDNAs encoding 1-deoxy-d-xylulose-5- phosphate synthase in Aquilaria sinensis (Lour.) Gilg. Plant Physiol. Biochem. 82:133-141. https://doi.org/10.1016/j.plaphy.2014.05.013
  41. Yang, L. -E., Huang, X. -Q., Lu, Q. -Q., Zhu, J. -Y. & Lu, S. 2016. Cloning and characterization of the geranylgeranyl diphosphate synthase (GGPS) responsible for carotenoid biosynthesis in Pyropia umbilicalis. J. Appl. Phycol. 28:671-678. https://doi.org/10.1007/s10811-015-0593-6
  42. Yang, L. -E., Jin, Q. -P., Xiao, Y., Xu, P. & Lu, S. 2013. Improved methods for basic molecular manipulation of the red alga Porphyra umbilicalis (Rhodophyta: Bangiales). J. Appl. Phycol. 25:245-252. https://doi.org/10.1007/s10811-012-9858-5
  43. Zhang, B. Y., Zhu, D. L., Wang, G. C. & Peng, G. 2014. Characterization of the AOX gene and cyanide-resistant respiration in Pyropia haitanensis (Rhodophyta). J. Appl. Phycol. 26:2425-2433. https://doi.org/10.1007/s10811-014-0274-x