Skip to main content
Applied Biological Chemistry
Download PDF

Expression of Jerusalem artichoke (Helianthus tuberosus L.) fructosyltransferases, and high fructan accumulation in potato tubers

Applied Biological Chemistry volume 62, Article number: 74 (2019) Cite this article

Abstract

Fructans are polymers of fructose that are present as storage carbohydrates in various plants. Jerusalem artichoke (Helianthus tuberosus L.) contains a high amount of inulin. Two enzymes are involved in inulin biosynthesis. The sucrose:sucrose 1-fructosyltransferase (1-SST) enzyme mainly catalyzes the synthesis of 1-kestose from sucrose. In the next step, fructan:fructan 1-fructosyltransferase (1-FFT) catalyzes the synthesis of inulin from 1-kestose. In this study, the Ht1-SST and Ht1-FFT genes were isolated from Jerusalem artichoke and expressed in potato (Solanum tuberosum L.), either separately or together, via Agrobacterium-mediated transformation. Transgenic potato tubers overexpressing Ht1-SST accumulated 1-kestose to a high level (up to 3.36 mg/g), while tubers overexpressing both Ht1-SST and Ht1-FFT accumulated up to 3.14 mg/g short-chain inulin-type fructans, with the degree of polymerization (DP) ranging from 3 to 5, excluding high DP inulins. Transgenic potato plants accumulated fructo-oligosaccharides to a high level, following the fructan biosynthetic pathway of Jerusalem artichoke, and therefore present a high potential for the mass production of inulin through established potato breeding and cultivation methods.

Introduction

Fructans are sucrose-derived water-soluble fructose polymers that commonly function as storage carbohydrates in more than 40,000 higher plant species including approximately 15% of flowering species, many belonging to the Poaceae, Liliaceae, and Asteraceae families [1]. Fructans are categorized into several groups, depending on their length (degree of polymerization [DP]), linkage type, and branching between fructose and glucose units [2]. The linear levan-type fructans consist of β-(2,6) linkages and are synthesized from 6-kestotriose in grasses such as Phleum pratense and Poa secunda [3, 4]. Graminan-type fructans consist of β-(1,2) and β-(2,6) linkages and are found in cereal crops such as wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) [5]. The most studied group of fructans is the linear inulin-type fructans. Inulins consist of linear β-(1,2) linkages with a terminal glucose residue and are present in Asteraceae family members such as chicory (Cichorium intybus L.), sunflower (Helianthus annuus L.), and Jerusalem artichoke (Helianthus tuberosus L.) [6]. Inulins are synthesized from sucrose by the action of two different fructosyltransferases, sucrose:sucrose 1-fructosyltransferase (1-SST; EC 2.4.1.99) and fructan:fructan 1-fructosyltransferase (1-FFT; EC 2.4.1.100) [7]. The 1-SST enzyme catalyzes the biosynthesis of the trisaccharide 1-kestotriose (DP3) from a sucrose molecule, while 1-FFT catalyzes the elongation of 1-kestotriose to inulins containing additional β-(1,2)-linked fructose units with higher DP [6, 8]. On the other hand, 6G-fructosyltransferase (6G-FFT; EC 2.4.1.243) is a key enzyme that catalyzes the synthesis of the inulin neoseries using 1-kestose as a fructose donor [9]. Recently, 6G-FFT was isolated from ryegrass (Lolium perenne L.) and functionally characterized [2]. Lp6G-FFT and native onion (Allium cepa L.) 6G-FFT exhibit 1-FFT activity and other similar properties [2, 10].

Jerusalem artichoke and chicory are the major industrial sources of inulin [11]. Jerusalem artichoke tubers contain a large amount of inulin and are therefore a valuable biomaterial for inulin production; 85% of the tuber dry weight corresponds to inulin-type fructans, with an intermediate DP (3–35 fructosyl residues) [6, 12]. Transgenic petunia (Petunia hybrida Vilm) plants overexpressing the Ht1-SST gene show the accumulation of low DP fructans, and the F1 progeny of a cross between two transgenic lines overexpressing Ht1-SST and Ht1-FFT show increased fructan content [13]. The chain length distribution of inulin-type fructans for globe artichoke (Cynara scolymus) and Jerusalem artichoke show no differences in the transient tobacco protoplast system [14, 15] as well as in transgenic maize (Zea mays L.), sugar beet (Beta vulgaris L.), and potato plants (Solanum tuberosum L.) [16, 17].

Inulin is a soluble dietary fiber with many health benefits. The addition of inulin in the diet helps to decrease lipogenesis in the liver [18], and oligofructose-enriched inulin prebiotics affect calcium absorption and bone mineralization [19]. Inulin-type prebiotics have also been reported to affect diarrhea and cell propagation in piglets [20]. Furthermore, inulin is used as a biomaterial for syrup and bioethanol production [21]. The successful industrialization of inulin products depends on cost-effective mass production technologies. Although inulin is abundant in Jerusalem artichoke tubers, its production is limited by the high cost. Transgenic crops may have a substantial advantage for inulin production worldwide using established breeding and cultivation protocols [17].

Potato is widely used for food and industrial purposes. We envision that potato plants engineered to produce inulin-type fructans will contribute to a broad spectrum of commodities in both the agricultural and industrial sectors. Here, we successfully induce overexpression of the Ht1-SST and Ht1-FFT genes in potato and confirm the production of inulin-type fructans in transgenic potato plants.

Materials and methods

Isolation of Ht1-SST and Ht1-FFT genes

Total RNA was isolated from the tubers of Jerusalem artichoke (Helianthus tuberosus cv. PJA) using the acid guanidinium thiocyanate-phenol–chloroform extraction method [22, 23]. First-strand cDNA was synthesized from 1 μg total RNA using AccuPower CycleScript RT PreMix (dT20) (Bioneer Co., Daejeon, Korea). Full-length Ht1-SST cDNA (1893 bp; GenBank accession number: AJ009757) was amplified from the cDNA template by PCR using primers HtSST_F (5′-AAAACCCTCCCTCAGGCCAC-3′) and HtSST_R (5′-CCATCAAAGTTCGAAAGTCC-3′) with the Expand Long PCR Kit (Roche Applied Science, Mannheim, Germany). Full-length Ht1-FFT cDNA (1848 bp; GenBank accession number: AJ009756.1) was amplified by PCR using primers HtFFT_F (5′-GTCAGTCACCATGCAAACCC-3′) and HtFFT_R (5′-AAAGGATAGCGATAGCCGG-3′). After sequence verification, the Ht1-SST and Ht1-FFT PCR products were cloned into the pCR8/GW/TOPO vector containing attL recombination sites using the pCR8/GW/TOPO TA Cloning Kit (Invitrogen, Carlsbad, CA), generating the Gateway entry clones of Ht1-SST and Ht1-FFT (Fig. 1a). The pEarleyGate103 (pEG103) vector containing the bar gene (as a selectable marker), the green fluorescent protein (GFP) reporter gene, and a 6x-His-tag was used as the Gateway destination vector containing attR recombination sites to generate pEG103/1-SST and pEG103/1-FFT expression constructs. These constructs were introduced into Agrobacterium tumefaciens strain GV3101, as described previously [24], which were then used for potato transformation.

Fig. 1
figure 1

Schematic representation of the vector construct and potato transformation process. a The Ht1-SST and Ht1-FFT genes were inserted in the pEG103 Gateway vector containing the cauliflower mosaic virus 35S promoter and bar gene. Each vector was introduced in wild-type potato plants. b Simplified diagram showing plant transformation and regeneration

Full size image

Plant material and transformation

Potato (Solanum tuberosum L. cv. Desiree) plants were cultivated in an incubation room at 24 °C, 16 h light/8 h dark photoperiod, and 50% relative humidity using aseptic technique. Stable potato transformation was performed by the cocultivation of leaf disc segments with Agrobacterium carrying the pEG103/1-SST and pEG103/1-FFT expression constructs, as described previously [25] (Fig. 1b). Transgenic shoots were induced on Murashige and Skoog (MS) medium containing 2.0 mg/L zeatin, 0.01 mg/L naphthalene acetic acid (NAA), 0.1 mg/L gibberellin (GA3), 500 mg/L carbenicillin, and 1.0 mg/L Basta. Roots were induced on MS medium containing 1.0 mg/L Basta and 500 mg/L carbenicillin for the selection of transgenic plants. Positive transformants displaying Ht1-SST activity were selected, and the Ht1-FFT gene was introduced in Ht1-SST transgenic potato plants to generate transgenic plants co-expressing Ht1-SST and Ht1-FFT (hereafter referred to as SnF plants). SnF transgenic potato plants expressing Ht1-SST and/or Ht1-FFT genes were selected based on antibiotic resistance, and then selected primarily by genomic DNA PCR using Plant Direct PCR system (NanoHelix Co., Daejeon, Korea) with full length SST and/or full length FFT primers. Expression of SST and/or FFT genes of transgenic plants at the RNA level was confirmed using RT-PCR and quantitative real-time PCR (qRT-PCR), and transgenic plant lines were selected.

Total RNA isolation and cDNA synthesis was performed as described previously [26]. All gene-specific primers were designed based on cDNA sequences (Table 1). Relative mRNA expression levels of Ht1-SST and Ht1-FFT were determined by qRT-PCR on a CFX Connect Real-Time PCR System (Bio-Rad, Hercules, CA) using a SYBR Green Master Mix (Enzynomics Co., Daejeon, Korea). The actin gene was used as a reference for data normalization. SnF transgenic plants introduced FFT-1 from transgenic plant expressing 1-SST were identified by RT-PCR with full-length 1-SST and 1-FFT primers, and SnF transgenic plant was selected.

Table 1 Primers used for reverse transcription PCR and quantitative real-time PCR (qRT-PCR) of fructosyltransferases and actin
Full size table

Extraction and analysis of water-soluble carbohydrates

Five grams (fresh weight) of tissue was sampled from each harvested potato tuber using a cork borer and then homogenized in 30 ml of 20 mM sodium phosphate buffer (pH 7.0). To extract water-soluble carbohydrates, samples were incubated in a water bath at 85 °C for 30 min. After cooling to room temperature, the extracts were centrifuged at 10,000×g and analyzed by high performance liquid chromatography (HPLC). Quantitative analysis of saccharides was performed using Sugar KS-802 column (8.0 × 300; Showa Denko K.K., Tokyo, Japan) with HPLC grade water (mobile phase) at 0.5 ml/min flow rate and 65 °C temperature on an HPLC refractive index detector (RID) system (Agilent 1100 series; Agilent Technologies, Santa Clara, CA).

Starch staining

The staining of starch in tuber tissues were performed using I2/KI solution (2 g KI, 1 g I2, 300 ml H2O). Longitudinal sections of transgenic potato tubers were stained for 5 min, and washed twice with distilled water, and examined.

Statistical analysis

Data were statistically processed using Excel 2010. Error bars represent the standard deviation (SD) of three independent biological replicates. Differences between means were considered statistically significant at p < 0.05.

Results and discussion

The Ht1-SST and Ht1-FFT genes were introduced in potato plants via Agrobacterium-mediated transformation [27]. The gene isolation, vector construction, plant growth, and plant transformation methodologies are shown in Fig. 1. Among the ten antibiotic-resistant plants, two independent transgenic lines expressing either Ht1-SST (S4 and S5) or Ht1-FFT (F4 and F5) were identified by Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) and Reverse transcription polymerase chain reaction (RT-PCR), which were performed using both newly synthesized cDNA [26] and gene-specific primers (Fig. 2a). The SnF transgenic potato plants expressing both Ht1-SST and Ht1-FFT were obtained by the introduction of Ht1-FFT in the transgenic line S5, chosen for its high level of Ht1-SST mRNA, followed by PCR-based selection of the Ht1-FFT gene (Fig. 2b). The phenotype, growth, and tuber size of all five independent transgenic lines (S4, S5, F4, F5, and SnF1) were compared with wild-type (WT) potato plants (Fig. 3a). Additionally, results of the starch iodine test showed no significant differences in sucrose starch metabolism among tubers of the five transgenic potato plants (Fig. 3b). These results suggest that the introduction of Ht1-SST and Ht1-FFT in potato plants has no negative impact on the growth, tuber formation, and starch biosynthesis of potato.

Fig. 2
figure 2

Expression analysis of Ht1-SST and Ht1-FFT in transgenic potato tubers. a Relative expression levels of Ht1-SST and Ht1-FFT mRNAs in transgenic potato tubers using quantitative real-time PCR (qRT-PCR) and reverse transcription PCR. Expression data were normalized relative to actin mRNA. Melting temperatures are indicated. b Identification of Ht1-SST and Ht1-FFT in SnF1 transgenic potato tubers by reverse transcription PCR. WT, wild-type potato cv. Desiree (negative control); S4, S5 and S10, independent transgenic potato lines expressing Ht1-SST; F4, F5, F6 and F8, independent transgenic potato lines expressing Ht1-FFT; SnF1, transgenic potato line co-expressing Ht1-SST and Ht1-FFT

Full size image
Fig. 3
figure 3

Phenotypic characterization of transgenic potato tubers expressing Ht1-SST and/or Ht1-FFT. a Phenotype of transgenic potato leaves and tubers. b Analysis of starch in transgenic potato tubers using the iodine staining assay. WT, wild-type potato; S4 and S5, independent transgenic potato lines expressing Ht1-SST; F4 and F5, independent transgenic potato lines expressing Ht1-FFT; SnF1, transgenic potato line co-expressing Ht1-SST and Ht1-FFT

Full size image

The 1-SST and 1-FFT genes encode important enzymes involved in the biosynthesis of inulin-type fructans. In this study, we performed HPLC analyses to determine the pattern of inulin-type fructans stored in transgenic potato plants expressing Ht1-SST and/or Ht1-FFT (Fig. 4). Compared with WT potato tubers, S4 and S5 potato tubers contained 3.38 ± 0.50 and 3.34 ± 0.80 mg/g DP3 inulin-type fructans, respectively (Table 2). We previously reported the measurement of inulin-type fructans from Jerusalem artichoke tubers [28]. Jerusalem artichoke tubers are composed of more than 70% fructans, so very high amounts of inulin-type fructans (approximately 693.22 mg/g) were detected, although the inulin was measured at the dry weight of the Jerusalem artichoke tuber. However, this high-expression inulin assay pattern was difficult to compare more clear results such as fructans synthesis process in inulin-type fructans analysis of transgenic potato. Therefore, we used the Jerusalem artichoke callus as a positive control, which shows relatively low inulin-type fructans synthesis compared to the Jerusalem artichoke tuber. Interestingly, DP3 inulin-type fructans accumulated to higher levels in S4 and S5 tubers than in Jerusalem artichoke callus (2.34 ± 0.82 mg/g) and SnF1 tubers (1.66 ± 0.04 mg/g). Additionally, a small amount of DP4 inulin-type fructans was detected in S4 and S5 tubers. According to Van Der Meer et al. [13], fructans in Jerusalem artichoke comprise β-(1,2)-linked fructose polymers, with a DP of up to 30. In this study, Jerusalem artichoke callus accumulated short-chain (DP3) inulin-type fructans. This finding is consistent with the observation of Pontis [29], who showed that differences in the length of fructans are not only caused by taxonomic variation but also by environmental conditions. Short-chain inulin-type fructans (DP3 and DP4) were identified in transgenic S4 and S5 potato tubers, while WT potato plants were confirmed to have no inulin-type fructans. On the other hand, no inulin-type fructans were detected in F4 or F5 tubers. These results confirmed the finding of Van Der Meer et al. [13] that 1-SST is responsible for the synthesis of DP3 inulin-type fructans from sucrose, whereas 1-FFT is unable to synthesize fructans from sucrose in the absence of 1-SST. Our results were also in agreement with previous reports of Ht1-SST-mediated induction of DP3 inulin-type fructans in heterologous systems, including sugar beet [30], petunia [13], and maize [17]. The SnF1 tubers accumulated short-chain inulin-type fructans to levels similar to the Jerusalem artichoke callus. Although SnF1 tubers showed a lower level of DP3 inulin-type fructans (1.66 ± 0.04 mg/g) than S4 and S5 tubers, inulin-type fructans with DP = 4 (0.88 ± 0.15 mg/g), DP = 5 (0.60 ± 0.15 mg/g), and DP > 5 (1.11 ± 0.11 mg/g) were also detected in SnF1 tubers. When fructans with DP ≥ 3 are used as a substrate, inulin-type fructans with a higher DP (> 3) are accumulated by the action of 1-FFT. This result confirms the role of 1-FFT in the production of short-chain inulin-type fructans using 1-kestose in the fructan biosynthetic pathway. Over 70% of inulin-type fructans stored in Jerusalem artichoke tubers represent DP3–DP20 fructans [31, 32]. Species-specific differences in the DP of fructans have been described previously; for example, the DP of inulin-type fructans is higher in globe artichoke than in chicory. These differences are thought to reflect species-specific activities of 1-FFT; for example, 1-FFT derived from globe artichoke (Cs1-FFT) shows high affinity for long-chain inulin-type fructans as an acceptor substrate, whereas 1-FFT derived from chicory (Ci1-FFT) prefers short-chain (DP3) inulin-type fructans as an acceptor substrate [33, 34]. Additionally, storage conditions (prolonged storage and temperature, 18 °C, 4 °C and 4 °C under polypropylene film packing) may cause a substantial decrease in the average DP of inulin-type fructans because of depolymerization or degradation of higher molecular weight carbohydrates [35]. Depolymerization or hydrolysis of fructans is mainly caused by fructan exohydrolase, which sequentially releases fructose units from the fructan polymer, eventually leaving sucrose [36]. The 1-FFT enzyme also contributes to the de-polymerization of fructans under unfavorable conditions [36, 37]. Therefore, we speculate that the accumulation of low DP inulin-type fructans in transgenic potato tubers was caused by 1-FFT postharvest, despite its role in fructan biosynthesis during tuber formation and maturation.

Fig. 4
figure 4

HPLC analysis of soluble carbohydrates extracted from transgenic and wild-type potato tubers. B, phosphate buffer; S, sucrose; DP3, 1-kestose; WT, wild-type potato; S4 and S5, independent transgenic potato lines expressing Ht1-SST; F4 and F5, independent transgenic potato lines expressing Ht1-FFT; SnF1, transgenic potato line co-expressing Ht1-SST and Ht1-FFT

Full size image
Table 2 Carbohydrate content of Jerusalem artichoke (Helianthus tuberosus L.) callus and transgenic potato tubers expressing Ht1-SST and/or Ht1-FFT
Full size table

Based on the activity of two enzymes, 1-SST and 1-FFT, fructan biosynthesis in another transgenic potato tuber has been previously reported. Transgenic potato tubers expressing 1-SST alone accumulated 1.42 mg/g DP3 inulin-type fructans, whereas those co-expressing 1-SST and 1-FFT accumulated 2.58 mg/g DP3–DP5 inulin-type fructans [15, 17]. In this study, the S4 and S5 transgenic tubers showed an average accumulation of 3.38 and 3.34 mg/g DP3 inulin-type fructans, respectively, which is approximately 2.4-fold greater than that achieved previously (Table 2). More specifically, two independent transgenic potato lines expressing Ht1-SST accumulated 3.67 and 3.61 mg/g DP3 and DP4 inulin-type fructans, respectively. In SnF1 transgenic tubers, short-chain (DP3–DP5) inulin-type fructans accumulated to 3.14 mg/g, while inulin-type fructans with DP > 5 accumulated to 1.11 mg/g. Overall, SnF1 tubers accumulated a total of approximately 4.25 mg/g inulin-type fructans; however, this value was less than the total amount of inulin-type fructans accumulated in Jerusalem artichoke callus (approximately 10.87 mg/g). We also compared the level of inulin-type fructans with DP ≥ 3 between S4 and SnF1 tubers (Fig. 4). While DP3 fructans accumulated to lower levels in SnF1 tubers than in S4 tubers, the level of fructans with DP > 3 was fivefold higher in SnF1 tubers (Fig. 4). These results demonstrate that in potato, 1-FFT produces inulin-type fructans using DP3 as a substrate (synthesized by 1-SST), although 1-SST uses DP3 to produce only a small amount of DP4 fructans. Sucrose synthesized in plants is transported to amyloplasts by sucrose transporters and stored as starch. Sucrose is also transferred to vacuoles [38]. Since fructan is synthesized and stored in the vacuole, inulin could be synthesized and stored in the vacuole by Ht1-SST and Ht1-FFT in transgenic potatoes. Overall, Ht1-SST and Ht1-FFT were successfully expressed in potato, either alone or in combination, resulting in the production of inulin-type fructans to higher levels than previously achieved. A brief schematic illustrates the successful accumulation of inulin-type fructans in the vacuole and the synthesis of starch in the amyloplast in transgenic potato tubers (Fig. 5). Fructan were also detected from some of the angiosperms adapted to region having cold and dry seasons [7]. It contribute to have resistance against abiotic stress by cold [39], drought [40, 41]. The improvement of stress tolerance by triggering fructan accumulation has been demonstrated in rice [42], potato [43], tobacco [44]. These transgenic potatoes could potentially be used as a nutritional supplement with health promoting properties, and/or as a main crop capable of withstanding relatively harsh cultivation conditions.

Fig. 5
figure 5

Schematic representation of expected metabolic pathway of inulin biosynthesis in transgenic potato tuber

Full size image

Availability of data and materials

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

References

  1. Hendry GA, Wallace RK (1993) The origin, distribution, and evolutionary significance of fructans. In: Suzuki M, Chatterton NJ (eds) Science and technology of fructans, pp 119–139

  2. Lasseur B, Lothier J, Djoumad A, De Coninck B, Smeekens S, Van Laere A et al (2006) Molecular and functional characterization of a cDNA encoding fructan: fructan 6G-fructosyltransferase (6G-FFT)/fructan: fructan 1-fructosyltransferase (1-FFT) from perennial ryegrass (Lolium perenne L.). J Exp Bot. 57(11):2719–2734

    Article  CAS  PubMed  Google Scholar 

  3. Chatterton N, Harrison P (1997) Fructan oligomers in Poa ampla. New Phytol 136(1):3–10

    Article  CAS  Google Scholar 

  4. Tamura K-I, Kawakami A, Sanada Y, Tase K, Komatsu T, Yoshida M (2009) Cloning and functional analysis of a fructosyltransferase cDNA for synthesis of highly polymerized levans in timothy (Phleum pratense L.). J Exp Bot. 60(3):893–905

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Van den Ende W, Clerens S, Vergauwen R, Van Riet L, Van Laere A, Yoshida M et al (2003) Fructan 1-exohydrolases. β-(2,1)-trimmers during graminan biosynthesis in stems of wheat? Purification, characterization, mass mapping, and cloning of two fructan 1-exohydrolase isoforms. Plant Physiol. 131(2):621–631

    Article  PubMed Central  CAS  Google Scholar 

  6. Edelman J, Jeeeord T (1968) The mechanisim of fructosan metabolism in higher plants as exemplified in Helianthus tuberosus. New Phytol 67(3):517–531

    Article  CAS  Google Scholar 

  7. Vijn I, Smeekens S (1999) Fructan: more than a reserve carbohydrate? Plant Physiol 120(2):351–360

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Van den Ende W, Van Laere A (1996) De novo synthesis of fructans from sucrose in vitro by a combination of two purified enzymes (sucrose: sucrose 1-fructosyl transferase and fructan: fructan 1-fructosyl transferase) from chicory roots (Cichorium intybus L.). Planta. 200(3):335–342

    Article  Google Scholar 

  9. Vijn I, Van Dijken A, Sprenger N, Van Dun K, Weisbeek P, Wiemken A et al (1997) Fructan of the inulin neoseries is synthesized in transgenic chicory plants (Cichorium intybus L.) harbouring onion (Allium cepa L.) fructan: fructan 6G-fructosyltransferase. Plant J. 11(3):387–398

    Article  CAS  PubMed  Google Scholar 

  10. Fujishima M, Sakai H, Ueno K, Takahashi N, Onodera S, Benkeblia N et al (2005) Purification and characterization of a fructosyltransferase from onion bulbs and its key role in the synthesis of fructo-oligosaccharides in vivo. New Phytol 165(2):513–524

    Article  CAS  PubMed  Google Scholar 

  11. Lingyun W, Jianhua W, Xiaodong Z, Da T, Yalin Y, Chenggang C et al (2007) Studies on the extracting technical conditions of inulin from Jerusalem artichoke tubers. J Food Eng 79(3):1087–1093

    Article  CAS  Google Scholar 

  12. Marx SP, Nösberger J, Frehner M (1997) Seasonal variation of fructan-β-fructosidase (FEH) activity and characterization of a β-(2–1)-linkage specific FEH from tubers of Jerusalem artichoke (Helianthus tuberosus). New Phytol 135(2):267–277

    Article  CAS  Google Scholar 

  13. Van Der Meer IM, Koops AJ, Hakkert JC, Van Tunen AJ (1998) Cloning of the fructan biosynthesis pathway of Jerusalem artichoke. Plant J 15(4):489–500

    Article  PubMed  Google Scholar 

  14. Hellwege EM, Raap M, Gritscher D, Willmitzer L, Heyer AG (1998) Differences in chain length distribution of inulin from Cynara scolymus and Helianthus tuberosus are reflected in a transient plant expression system using the respective 1-FFT cDNAs. FEBS Lett 427(1):25–28

    Article  CAS  PubMed  Google Scholar 

  15. Hellwege EM, Czapla S, Jahnke A, Willmitzer L, Heyer AG (2000) Transgenic potato (Solanum tuberosum) tubers synthesize the full spectrum of inulin molecules naturally occurring in globe artichoke (Cynara scolymus) roots. Proc Natl Acad Sci 97(15):8699–8704

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Koops AJ, Sevenier R, Van Tunen AJ, De Leenheer L (2003) Transgenic plants presenting a modified inulin producing profile. Google Patents; US 6664444 B1

  17. Stoop JM, Van Arkel J, Hakkert JC, Tyree C, Caimi PG, Koops AJ (2007) Developmental modulation of inulin accumulation in storage organs of transgenic maize and transgenic potato. Plant Sci 173(2):172–181

    Article  CAS  Google Scholar 

  18. Delzenne NM, Kok N (2001) Effects of fructans-type prebiotics on lipid metabolism. Am J Clin Nutr. 73(2):456s–458s

    Article  CAS  PubMed  Google Scholar 

  19. Franck A (2006) Oligofructose-enriched inulin stimulates calcium absorption and bone mineralisation. Nutr Bull. 31(4):341–345

    Article  Google Scholar 

  20. Kien CL, Chang J, Cooper JR, Frankel WL (2004) Effects of prefeeding a prebiotic on diarrhea and colonic cell proliferation in piglets fed lactulose. J Par Enter Nutr. 28(1):22–26

    Article  CAS  Google Scholar 

  21. Chi Z, Chi Z, Zhang T, Liu G, Yue L (2009) Inulinase-expressing microorganisms and applications of inulinases. Appl Microbiol Biotechnol 82(2):211–220

    Article  CAS  PubMed  Google Scholar 

  22. Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162(1):156–159

    Article  CAS  PubMed  Google Scholar 

  23. Chomczynski P, Sacchi N (2006) The single-step method of RNA isolation by acid guanidinium thiocyanate–phenol–chloroform extraction: twenty-something years on. Nat Protoc 1(2):581–585

    Article  CAS  PubMed  Google Scholar 

  24. Holsters M, De Waele D, Depicker A, Messens E, Van Montagu M, Schell J (1978) Transfection and transformation of Agrobacterium tumefaciens. Mol Gen Genet 163(2):181–187

    Article  CAS  PubMed  Google Scholar 

  25. Kim H-S, Euym J-W, Kim M-S, Lee B-C, Mook-Jung I, Jeon J-H et al (2003) Expression of human β-amyloid peptide in transgenic potato. Plant Sci 165(6):1445–1451

    Article  CAS  Google Scholar 

  26. Moon K-B, Lee J, Kang S, Kim M, Mason HS, Jeon J-H et al (2014) Overexpression and self-assembly of virus-like particles in Nicotiana benthamiana by a single-vector DNA replicon system. Appl Microbiol Biotechnol 98(19):8281–8290

    Article  CAS  PubMed  Google Scholar 

  27. Kim M-J, An D-J, Moon K-B, Cho H-S, Min S-R, Sohn J-H et al (2016) Highly efficient plant regeneration and Agrobacterium-mediated transformation of Helianthus tuberosus L. Ind Crops Prod 83:670–679

    Article  CAS  Google Scholar 

  28. Min CW, Jung WY, Park HJ, Moon K-B, Ko H, Sohn J-H et al (2019) Label-free quantitative proteomic analysis determines changes in amino acid and carbohydrate metabolism in three cultivars of Jerusalem artichoke tubers. Plant Biotechnol Rep. 13(2):111–122

    Article  Google Scholar 

  29. Pontis HG (1989) Fructans and cold stress. J Plant Physiol 134(2):148–150

    Article  CAS  Google Scholar 

  30. Weyens G, Ritsema T, Van Dun K, Meyer D, Lommel M, Lathouwers J et al (2004) Production of tailor-made fructans in sugar beet by expression of onion fructosyltransferase genes. Plant Biotechnol J 2(4):321–327

    Article  CAS  PubMed  Google Scholar 

  31. Praznik W, Beck R (1985) Application of gel permeation chromatographic systems to the determination of the molecular weight of inulin. J Chromatogr A 348:187–197

    Article  CAS  Google Scholar 

  32. Srinameb B-O, Nuchadomrong S, Jogloy S, Patanothai A, Srijaranai S (2015) Preparation of Inulin Powder from Jerusalem Artichoke (Helianthus tuberosus L.) Tuber. Plant Foods Hum Nutr. 70(2):221–226

    Article  CAS  PubMed  Google Scholar 

  33. Koops AJ, Jonker HH (1994) Purification and characterization of the enzymes of fructan biosynthesis in tubers of Helianthus tuberosus ‘Colombia’I. Fructan: fructan fructosyl transferase. J Exp Bot. 45(11):1623–1631

    Article  CAS  Google Scholar 

  34. Vergauwen R, Van Laere A, Van den Ende W (2003) Properties of fructan: fructan 1-fructosyltransferases from chicory and globe thistle, two Asteracean plants storing greatly different types of inulin. Plant Physiol 133(1):391–401

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Leroy G, Grongnet JF, Mabeau S, Corre DL, Baty-Julien C (2010) Changes in inulin and soluble sugar concentration in artichokes (Cynara scolymus L.) during storage. J Sci Food Agric. 90(7):1203–1209

    Article  CAS  PubMed  Google Scholar 

  36. Xu H, Liang M, Xu L, Li H, Zhang X, Kang J et al (2015) Cloning and functional characterization of two abiotic stress-responsive Jerusalem artichoke (Helianthus tuberosus) fructan 1-exohydrolases (1-FEHs). Plant Mol Biol 87(1–2):81–98

    Article  CAS  PubMed  Google Scholar 

  37. Degasperi MI, Itaya NM, Buckeridge MS, Figueiredo-Ribeiro RDCL (2003) Fructan degradation and hydrolytic activity in tuberous roots of Viguiera discolor Baker (Asteraceae), a herbaceous species from the cerrado. Braz J Bot. 26(1):11–21

    Article  CAS  Google Scholar 

  38. Nookaraju A, Upadhyaya CP, Pandey SK, Young KE, Hong SJ, Park SK et al (2010) Molecular approaches for enhancing sweetness in fruits and vegetables. Sci Hortic 127(1):1–15

    Article  CAS  Google Scholar 

  39. Hisano H, Kanazawa A, Yoshida M, Humphreys MO, Iizuka M, Kitamura K et al (2008) Coordinated expression of functionally diverse fructosyltransferase genes is associated with fructan accumulation in response to low temperature in perennial ryegrass. New Phytol 178(4):766–780

    Article  CAS  PubMed  Google Scholar 

  40. De Roover J, Vandenbranden K, Van Laere A, Van den Ende W (2000) Drought induces fructan synthesis and 1-SST (sucrose:sucrose fructosyltransferase) in roots and leaves of chicory seedlings (Cichorium intybus L.). Planta. 210(5):808–814

    Article  PubMed  Google Scholar 

  41. Livingston DP, Hincha DK, Heyer AG (2009) Fructan and its relationship to abiotic stress tolerance in plants. Cell Mol Life Sci 66(13):2007–2023

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Kawakami A, Sato Y, Yoshida M (2008) Genetic engineering of rice capable of synthesizing fructans and enhancing chilling tolerance. J Exp Bot 59(4):793–802

    Article  CAS  PubMed  Google Scholar 

  43. Knipp G, Honermeier B (2006) Effect of water stress on proline accumulation of genetically modified potatoes (Solanum tuberosum L.) generating fructans. J Plant Physiol. 163(4):392–397

    Article  CAS  PubMed  Google Scholar 

  44. Li H-J, Yang A-F, Zhang X-C, Gao F, Zhang J-R (2007) Improving freezing tolerance of transgenic tobacco expressing sucrose: sucrose 1-fructosyltransferase gene from Lactuca sativa. Plant Cell Tissue Organ Cult 89(1):37–48

    Article  CAS  Google Scholar 

Download references

Funding

This work was supported by the Bio & Medical Technology Development Program of the National Research Foundation of Korea (No. 2015M3A9A5031107), funded by the Korean government.

Author information

Authors and Affiliations

  1. Plant Systems Engineering Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, South Korea

    Ki-Beom Moon, Ji-Sun Park, Hye-Sun Cho, Hyun-Soon Kim & Jae-Heung Jeon

  2. Department of Biological Sciences, Chungnam National University, Daejeon, 34148, South Korea

    Ki-Beom Moon & Youn-il Park

  3. Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, South Korea

    Hyunjun Ko & Jung-Hoon Sohn

Authors
  1. Ki-Beom Moon
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hyunjun Ko
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ji-Sun Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jung-Hoon Sohn
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hye-Sun Cho
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Youn-il Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hyun-Soon Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Jae-Heung Jeon
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

JHJ and HSK designed and coordinated the completion of the experiments. KB and JS grew and maintained plants. H-SK and JHS performed HPLC and analyzed the data. KB, HSC, YI, HSK and JHJ analyzed the entire data. KB, HSK and JHJ wrote the manuscript. HSC and YI provided guidance and supervised the study. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Hyun-Soon Kim or Jae-Heung Jeon.

Ethics declarations

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Moon, KB., Ko, H., Park, JS. et al. Expression of Jerusalem artichoke (Helianthus tuberosus L.) fructosyltransferases, and high fructan accumulation in potato tubers. Appl Biol Chem 62, 74 (2019). https://doi.org/10.1186/s13765-019-0481-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s13765-019-0481-x

Keywords