Issue
Molecular cloning and biotic elicitation response of phenylalanine ammonia-lyase gene of Astragalus chrysochlorus
Corresponding Author(s) : Neslihan Turgut Kara
Cellular and Molecular Biology,
Vol. 64 No. 5: Issue 5
Abstract
Phenylalanine ammonia lyase (PAL) is the first enzyme of the phenylpropanoid pathway, and it is necessary to upregulate flavonoid biosynthesis in most of the plant species. In this study, we have cloned PAL gene from endemic Astragalus chrysochlorus which is a producer of phenolic nicotiflorin (kaempferol-3-O-rutinoside). The cDNA encoding PAL was cloned from A. chrysochlorus using RT-PCR (reverse transcription-polymerase chain reaction) with conserved primer pairs. Amino acid sequence alignments showed that AcPAL (2160 bp, Accession number: KM189182) has more than 95% amino acid identity with their homologues in other Astragalus species. The coding sequence for the protein of AcPAL is 720 amino acids with a calculated molecular weight of 78.53 kDa. Full length AcPAL was cloned and expressed in Escherichia coli. qPCR (quantitative real-time PCR) analysis of the expression of PAL gene of A. chrysochlorus suggested that maximum transcript level was observed in 3 h yeast extract elicited suspension cells. Our findings suggest that AcPAL plays role in early response for yeast extract treatment. The isolation of AcPAL gene could be result in further studies for overproduction of secondary metabolite, nicotiflorin.
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- Hahlbrock K, Scheel D. Physiology and molecular-biology of phenylpropanoid metabolism. Annu Rev Plant Phys 1989; 40: 347-369.
- Boudet AM. Evolution and current status of research in phenolic compounds. Phytochemistry 2007; 68: 2722-2735.
- Miller AL. Botanical influences on cardiovascular disease. Altern Med Rev,3, 422-431.
- Sinclair S. Chinese herbs: A clinical review of astragalus, ligusticum, and schizandrae. Altern Med Rev 1998; 3: 338-344.
- Ma XQ, Shi Q, Duan JA, Dong TT, Tsim KW. Chemical analysis of radix astragali (huangqi) in china: A comparison with its adulterants and seasonal variations. J Agric Food Chem 2002; 50: 4861-4866.
- Xiao HB, Krucker M, Albert K, Liang XM. Determination and identification of isoflavonoids in radix astragali by matrix solid-phase dispersion extraction and high-performance liquid chromatography with photodiode array and mass spectrometric detection. J Chromatogr A 2004; 1032: 117-124.
- Aytaç Z. The revision of the section dasyphyllium bunge of the genus. Turk J Bot 1997; 31-57.
- Hasançebi S. Studies on tissue culture and transformation of Astragalus chrysochlorus. Istanbul University, PhD Thesis, Istanbul, 2003.
- Karagoz A, Turgut-Kara N, Cakir O, Demirgan R, Ari S. Cytotoxic activity of crude extracts from Astragalus chrysochlorus (leguminosae). Biotechnol Biotec Eq 2007; 21: 220-222.
- Ari S, Cakir O, Turgut-Kara N. Selenium tolerance in Astragalus chrysochlorus: Identification of a cdna fragment encoding a putative selenocysteine methyltransferase. Acta Physiol Plant 2010; 32: 1085-1092.
- Li RP, Guo ML, Zhang G, Xu XF, Li Q. Neuroprotection of nicotiflorin in permanent focal cerebral ischemia and in neuronal cultures. Biol Pharm Bull 2006; 29: 1868-1872.
- Lim SS, Jung YJ, Hyun SK, Lee YS, Choi JS. Rat lens aldose reductase inhibitory constituents of nelumbo nucifera stamens. Phytother Res 2006; 20; 825-830.
- Yang H-J, Myoung-Chong S, Myun-Ho B, Jin-Hee L, In-Sik C, Youn-Hyung L, Tae-Sook J, Byoung-Mog K, Sung-Hoon K, Dae-Keun K, Mi-Hyun P, Nam-In B. Development of biologically active compounds from edible plant sources-xii. Flavonol glycosides from trigonotis peduncularis benth and its hacat1 inhibitory activity. J Korean Soc Appl Biol Chem 2005; 48: 98-102.
- Akbay P, Basaran AA, Undeger U, Basaran N. In vitro immunomodulatory activity of flavonoid glycosides from urtica dioica l. Phytother Res 2003; 17: 34-37.
- Ishiguro K, Ohira Y, Oku H. Preventive effects of impatiens balsamina on the hen egg-white lysozyme (hel)-induced decrease in blood flow. Biol Pharm Bull 2002; 25: 505-508.
- Abreu PM, Braham H, Ben Jannet H, Mighri Z, Matthew S. Antioxidant compounds from ebenus pinnata. Fitoterapia 2007; 78: 32-34.
- Shyaula SL, Abbas G, Siddiqui H, Sattar SA, Choudhary MI, Basha FZ. Synthesis and antiglycation activity of kaempferol-3-o-rutinoside (nicotiflorin). Med Chem 2012; 8: 415-420.
- Turgut-Kara N, í‡akır í–, Hasançebi S, Karabey F, Arı Åž . Identification and production of phenolic nicotiflorin in Astragalus chrysochlorus callus. Farmacia 2017 (Accepted).
- Turgut-Kara N, Kahraman BU. Effects of long-term culture of astragalus chrysochlorus callus onmorphology, genetic structure, gene expression and metabolism. Plant Biosyst 2015; 149: 329-336.
- Cakir O, Ari S. Defensive and secondary metabolism in astragalus chrysochlorus cell cultures, in response to yeast extract stressor. J Environ Biol 2009; 30: 51-55.
- Murashige T, Skoog FA (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 1998; 15: 473-497.
- Chen H, Chen F. Effect of yeast elicitor on the secondary metabolism of ti-transformed salvia miltiorrhiza cell suspension cultures. Plant Cell Rep; 19: 710-717.
- Fredslund J, Schauser L, Madsen LH, Sandal N, Stougaard J. Prifi: Using a multiple alignment of related sequences to find primers for amplification of homologs. Nucleic Acids Res 2005; 33: W516-W520.
- Dixon RA, Paiva NL. Stress-induced phenylpropanoid metabolism. Plant Cell 1995; 7: 1085-1097.
- Liu RR, Xu SH, Li JL, Hu YL, Lin ZP. Expression profile of a pal gene from astragalus membranaceus var. Mongholicus and its crucial role in flux into flavonoid biosynthesis. Plant Cell Rep 2006; 25: 705-710.
- Hamerski D, Beier RC, Kneusel RE, Matern U, Himmelspach K. Accumulation of coumarins in elicitor-treated cell-suspension cultures of ammi-majus. Phytochemistry 1990; 29: 1137-1142.
References
Hahlbrock K, Scheel D. Physiology and molecular-biology of phenylpropanoid metabolism. Annu Rev Plant Phys 1989; 40: 347-369.
Boudet AM. Evolution and current status of research in phenolic compounds. Phytochemistry 2007; 68: 2722-2735.
Miller AL. Botanical influences on cardiovascular disease. Altern Med Rev,3, 422-431.
Sinclair S. Chinese herbs: A clinical review of astragalus, ligusticum, and schizandrae. Altern Med Rev 1998; 3: 338-344.
Ma XQ, Shi Q, Duan JA, Dong TT, Tsim KW. Chemical analysis of radix astragali (huangqi) in china: A comparison with its adulterants and seasonal variations. J Agric Food Chem 2002; 50: 4861-4866.
Xiao HB, Krucker M, Albert K, Liang XM. Determination and identification of isoflavonoids in radix astragali by matrix solid-phase dispersion extraction and high-performance liquid chromatography with photodiode array and mass spectrometric detection. J Chromatogr A 2004; 1032: 117-124.
Aytaç Z. The revision of the section dasyphyllium bunge of the genus. Turk J Bot 1997; 31-57.
Hasançebi S. Studies on tissue culture and transformation of Astragalus chrysochlorus. Istanbul University, PhD Thesis, Istanbul, 2003.
Karagoz A, Turgut-Kara N, Cakir O, Demirgan R, Ari S. Cytotoxic activity of crude extracts from Astragalus chrysochlorus (leguminosae). Biotechnol Biotec Eq 2007; 21: 220-222.
Ari S, Cakir O, Turgut-Kara N. Selenium tolerance in Astragalus chrysochlorus: Identification of a cdna fragment encoding a putative selenocysteine methyltransferase. Acta Physiol Plant 2010; 32: 1085-1092.
Li RP, Guo ML, Zhang G, Xu XF, Li Q. Neuroprotection of nicotiflorin in permanent focal cerebral ischemia and in neuronal cultures. Biol Pharm Bull 2006; 29: 1868-1872.
Lim SS, Jung YJ, Hyun SK, Lee YS, Choi JS. Rat lens aldose reductase inhibitory constituents of nelumbo nucifera stamens. Phytother Res 2006; 20; 825-830.
Yang H-J, Myoung-Chong S, Myun-Ho B, Jin-Hee L, In-Sik C, Youn-Hyung L, Tae-Sook J, Byoung-Mog K, Sung-Hoon K, Dae-Keun K, Mi-Hyun P, Nam-In B. Development of biologically active compounds from edible plant sources-xii. Flavonol glycosides from trigonotis peduncularis benth and its hacat1 inhibitory activity. J Korean Soc Appl Biol Chem 2005; 48: 98-102.
Akbay P, Basaran AA, Undeger U, Basaran N. In vitro immunomodulatory activity of flavonoid glycosides from urtica dioica l. Phytother Res 2003; 17: 34-37.
Ishiguro K, Ohira Y, Oku H. Preventive effects of impatiens balsamina on the hen egg-white lysozyme (hel)-induced decrease in blood flow. Biol Pharm Bull 2002; 25: 505-508.
Abreu PM, Braham H, Ben Jannet H, Mighri Z, Matthew S. Antioxidant compounds from ebenus pinnata. Fitoterapia 2007; 78: 32-34.
Shyaula SL, Abbas G, Siddiqui H, Sattar SA, Choudhary MI, Basha FZ. Synthesis and antiglycation activity of kaempferol-3-o-rutinoside (nicotiflorin). Med Chem 2012; 8: 415-420.
Turgut-Kara N, í‡akır í–, Hasançebi S, Karabey F, Arı Åž . Identification and production of phenolic nicotiflorin in Astragalus chrysochlorus callus. Farmacia 2017 (Accepted).
Turgut-Kara N, Kahraman BU. Effects of long-term culture of astragalus chrysochlorus callus onmorphology, genetic structure, gene expression and metabolism. Plant Biosyst 2015; 149: 329-336.
Cakir O, Ari S. Defensive and secondary metabolism in astragalus chrysochlorus cell cultures, in response to yeast extract stressor. J Environ Biol 2009; 30: 51-55.
Murashige T, Skoog FA (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 1998; 15: 473-497.
Chen H, Chen F. Effect of yeast elicitor on the secondary metabolism of ti-transformed salvia miltiorrhiza cell suspension cultures. Plant Cell Rep; 19: 710-717.
Fredslund J, Schauser L, Madsen LH, Sandal N, Stougaard J. Prifi: Using a multiple alignment of related sequences to find primers for amplification of homologs. Nucleic Acids Res 2005; 33: W516-W520.
Dixon RA, Paiva NL. Stress-induced phenylpropanoid metabolism. Plant Cell 1995; 7: 1085-1097.
Liu RR, Xu SH, Li JL, Hu YL, Lin ZP. Expression profile of a pal gene from astragalus membranaceus var. Mongholicus and its crucial role in flux into flavonoid biosynthesis. Plant Cell Rep 2006; 25: 705-710.
Hamerski D, Beier RC, Kneusel RE, Matern U, Himmelspach K. Accumulation of coumarins in elicitor-treated cell-suspension cultures of ammi-majus. Phytochemistry 1990; 29: 1137-1142.
