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Establishment and biochemical characterization of tamarillo (Solanum betaceum Cav.) embryogenic cell suspension cultures

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Abstract

Somatic embryogenesis (SE) is an important biotechnological tool with great potential for large-scale cloning. In Solanum betaceum Cav. (tamarillo), embryogenic (EC) and non-embryogenic (non-EC) cells can be obtained from the same explant on auxin-containing medium, making this system ideal for the evaluation of biochemical changes occurring during embryogenic induction. Liquid cultures offer additional possibilities for the analysis of factors controlling SE induction, and the main objectives here were the establishment of cell suspensions and the characterization of the extracellular protein profiles in EC and non-EC cultures. Growth kinetics of liquid cultures, starting with different amounts of EC or non-EC callus or with different weight per volume ratios, were analyzed. Embryogenic suspension cultures were efficiently established starting with 40 mg of cells in 20 mL of liquid medium. Mass spectrometry and fluorometric techniques were employed to identify extracellular proteins, their hydrolytic activity, and the main classes of proteases secreted into the media of EC or non-EC cultures. Extracellular protein profiles revealed quantitative and qualitative differences between EC and non-EC suspension cultures, mainly for several hydrolytic enzymes, such as glucanases and xylanases. Proteolytic activity analysis found serine proteases, aspartic proteases, and metalloproteases in EC cultures, whereas serine proteases were dominant in non-EC lines. For the first time, a protocol for the growth of tamarillo EC and non-EC suspensions was achieved. Moreover, the comparison of protein profiles between EC and non-EC lines showed pronounced differences in the proteolytic and glycolytic enzymes secreted.

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References

  • Abid G, Muhovski Y, Jacquemin JM, Mingeot D, Sassi K, Toussaint A, Baudoin J-P (2011) In silico identification and characterization of putative differentially expressed genes involved in common bean (Phaseolus vulgaris L.) seed development. Plant Cell Tissue Organ Cult 107:341–353

    Article  CAS  Google Scholar 

  • Aitken A, Learmonth MP (2002) Protein determination by UV absorption. In: Walker JM (ed) The protein protocols handbook. Humana Press, New York, pp 3–6

    Chapter  Google Scholar 

  • Blewett J, Burrows K, Thomas C (2000) A micromanipulation method to measure the mechanical properties of single tomato suspension cells. Biotechnol Lett 22:1877–1883

    Article  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Brown MB, Forsythe AB (1974) The ANOVA and multiple comparisons for data with heterogeneous variances. Biometrics 30:719–724

    Article  Google Scholar 

  • Canhoto JM, Lopes ML, Cruz GS (2005) Protocol of somatic embryogenesis: tamarillo (Cyphomandra betacea (Cav.) Sendtn.) In: Jain SM, Gupta PK (eds) Protocol for somatic embryogenesis in woody plants. Springer, Netherlands, pp 379–389

    Chapter  Google Scholar 

  • Carlberg I, Jonsson L, Bergenstråhle A, Söderhäll K (1987) Purification of a trypsin inhibitor secreted by embryogenic carrot cells. Plant Physiol 84:197–290

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Carpita NC, Gibeaut DM (1993) Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth. Plant J 3:1–30

    Article  CAS  PubMed  Google Scholar 

  • Catalá C, Rose JKC, Bennett AB (2000) Auxin-regulated genes encoding cell wall-modifying proteins are expressed during early tomato fruit growth. Plant Physiol 122:527–534

    Article  PubMed  PubMed Central  Google Scholar 

  • Catalá C, Rose JKC, York WS, Albersheim P, Darvill AG, Bennett AB (2001) Characterization of a tomato xyloglucan endotransglycosylase gene that is down-regulated by auxin in etiolated hypocotyls. Plant Physiol 127:1180–1192

    Article  PubMed  PubMed Central  Google Scholar 

  • Chawla HS (2010) Introduction to plant biotechnology, 3rd edn. Science Publishers Inc, Enfield New Hampshire

    Google Scholar 

  • Chen X-Y, Kim ST, Cho WK, Rim Y, Kim S, Kim S-W, Kang KY, Park ZY, Kim J-Y (2009) Proteomics of weakly bound cell wall proteins in rice calli. J Plant Physiol 166:675–685

    Article  CAS  PubMed  Google Scholar 

  • Chitteti BR, Tan F, Mujahid H, Magee BG, Bridges SM, Peng Z (2008) Comparative analysis of proteome differential regulation during cell dedifferentiation in Arabidopsis. Proteomics 8:4303–4316

    Article  CAS  PubMed  Google Scholar 

  • Chong-Pérez B, Reyes M, Rojas L, Ocaña B, Pérez B, Kosky RG, Angenon G (2012) Establishment of embryogenic cell suspension cultures and Agrobacterium-mediated transformation in banana cv. ‘Dwarf Cavendish’ (Musa AAA): effect of spermidine on transformation efficiency. Plant Cell Tissue Organ Cult 111:79–90

    Article  Google Scholar 

  • Chugh A, Khurana P (2002) Gene expression during somatic embryogenesis—recent advances. Curr Sci 86:715–730

    Google Scholar 

  • Correia SI, Canhoto JM (2012) Biotechnology of tamarillo (Cyphomandra betacea): from in vitro cloning to genetic transformation. Sci Hortic (Amsterdam) 148:161–168

    Article  CAS  Google Scholar 

  • Correia SI, Lopes ML, Canhoto JM (2011) Somatic embryogenesis induction system for cloning an adult Cyphomandra betacea (Cav.) Sendt. (tamarillo). Trees 25:1009–1020

    Article  Google Scholar 

  • Correia S, Vinhas R, Manadas B, Lourenço AS, Veríssimo P, Canhoto JM (2012) Comparative proteomic analysis of auxin-induced embryogenic and non embryogenic tissues of the solanaceous tree Chyphomandra batacea (tamarillo). J Proteome Res 11:1666–1675

    Article  CAS  PubMed  Google Scholar 

  • Correia SI, Alves AC, Veríssimo P, Canhoto JM (2015) Somatic embryogenesis in broad-leaf woody plants: what we can learn from proteomics. In: Germanà MA, Lambardi M (eds) In vitro embryogenesis in higher plants, Methods in molecular biology, vol 1359. Springer Science+Business Media, New York, pp 117–130

    Chapter  Google Scholar 

  • Cosgrove DJ (2005) Growth of the plant cell wall. Nat Rev Mol Cell Biol 6:850–861

    Article  CAS  PubMed  Google Scholar 

  • de Klerk G-J, ter Brugge J (2010) Micropropagation of Alstroemeria in liquid medium using slow release of medium components. Propag Ornam Plants 10:246–252

    Google Scholar 

  • de Vries SC, Booij H, Meyerink P, Huisman G, Wilde HD, Thomas TL, van Kammen A (1988) Acquisition of embryogenic potential in carrot cell-suspension cultures. Planta 176:196–204

    Article  PubMed  Google Scholar 

  • Fehér A (2015) Somatic embryogenesis—stress-induced remodelling of plant cell fate. Biochim Biophys Acta 1849:385–402

    Article  PubMed  Google Scholar 

  • Fehér A, Pasternak TP, Dudits D (2003) Transition of somatic plant cells to an embryogenic state. Plant Cell Tissue Organ Cult 74:201–228

    Article  Google Scholar 

  • Guimarães ML, Tomé MC, Crux GS (1996) Cyphomandra betacea (Cav.) Sendtn. (tamarillo). In YPS Bajaj Biotechnology in agriculture and forestry 35 Trees IV. Springer-Verlag Berlin Heidelberg New York pp 120–136

  • Hurtado NH, Morales AL, González-Miret ML, Escudero-Gilete ML, Heredia FJ (2009) Colour, pH stability and antioxidant activity of anthocyanin rutinoside isolated from tamarillo fruit (Solanum betaceum Cav). Food Chem 117:88–93

    Article  CAS  Google Scholar 

  • Jamet E, Albenne C, Boudart G, Irshad M, Canut H, Pont-Lezica R (2008) Recent advances in plant cell wall proteomics. Proteomics 8:893–908

    Article  CAS  PubMed  Google Scholar 

  • Komamine A, Murata N, Nomura K (2005) Mechanisms of somatic embryogenesis in carrot suspension cultures—morphology, physiology, biochemistry, and molecular biology. In Vitro Cell Dev Biol – Plant 41:6–10

    Article  CAS  Google Scholar 

  • Kou M, Yen J, Hong J, Wang C (2008) Cyphomandra betacea Sendt. phenolics protect LDL from oxidation and PC12 cells from oxidative stress. LWT – Food Sci Technol 42:458–463

    Article  Google Scholar 

  • Lopes ML, Ferreira MR, Carloto JM, Cruz GS, Canhoto JM (2000) Somatic embryogenesis induction in tamarillo (Cyphomandra betacea). In: Jain SM, Gupta PK, Newton RJ (eds) Somatic embryogenesis in woody plants, vol 6. Kluwer Academic Publishers, Dordrecht, pp 433–455

    Chapter  Google Scholar 

  • Maës O, Coutos-Thévenot P, Jouenne T, Boulay M, Guern J (1997) Influence of extracellular proteins, proteases and protease inhibitors on grapevine somatic embryogenesis. Plant Cell Tissue Organ Cult 50:97–105

    Article  Google Scholar 

  • Malinowski R, Filipecki M, Tagashira N, Wisniewska A, Gaj P, Plader W, Malepzy S (2004) Xyloglucan endotransglucosylase/hydrolase genes in cucumber (Cucumis sativus)–differential expression during somatic embryogenesis. Physiol Plant 120:678–685

    Article  CAS  PubMed  Google Scholar 

  • Maloney VJ, Samuels AL, Mansfield SD (2012) The endo-1,4-beta-glucanase Korrigan exhibits functional conservation between gymnosperms and angiosperms and is required for proper cell wall formation in gymnosperms. New Phytol 193:1076–1087

    Article  CAS  PubMed  Google Scholar 

  • Marsoni M, Bracale M, Espen L, Prinsi B, Negri A, Vannini C (2008) Proteomic analysis of somatic embryogenesis in Vitis vinifera. Plant Cell Rep 27:347–356

    Article  CAS  PubMed  Google Scholar 

  • Mitsuhashi W, Yamashita T, Toyomasu T, Kashiwagi Y, Konnai T (2004) Sequential development of cysteine proteinase activities and gene expression during somatic embryogenesis in carrot. Biosci Biotechnol Biochem 68:705–713

    Article  CAS  PubMed  Google Scholar 

  • Morton JF (1987) Tree tomato. In: Morton JF (ed) Fruits of warm climates, Miami FL, pp 437–440

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  • Mustafa NR, de Winter W, van Iren F, Verpoorte R (2011) Initiation, growth and cryopreservation of plant cell suspension cultures. Nat Protoc 6:715–742

    Article  CAS  PubMed  Google Scholar 

  • Nicol F, His I, Jauneau A, Vernhettes S, Canut H, Höfte H (1998) A plasma membrane-bound putative endo-1,4-beta-D-glucanase is required for normal wall assembly and cell elongation in Arabidopsis. EMBO J 17:5563–5576

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Niemenak N, Awah TM, Lieberei R (2012) Establishment of suspension culture in Theobroma cacao and polyamines associated with cacao embryogenesis. Plant Growth Regul 67:1–8

    Article  CAS  Google Scholar 

  • Oropeza M, Marcano AK, García ED (2001) Proteins related with embryogenic potential in callus and cell suspensions of sugarcane (Saccharum sp.) In Vitro Cell Dev Biol – Plant 37:211–216

    Article  CAS  Google Scholar 

  • Pavoković D, Poljuha D, Horvatić A, Ljubešić N, Hagège D, Krsnik-Rasol M (2012) Morphological and proteomic analyses of sugar beet cultures and identifying putative markers for cell differentiation. Plant Cell Tissue Organ Cult 108:111–119

    Article  Google Scholar 

  • Prohens J, Nuez F (2010) The tamarillo (Cyphomandra betacea). Small Fruits Review 1:43–68

    Article  Google Scholar 

  • Qin Q, Bergmann CW, Rose JK, Saladie M, Kolli VS, Albersheim P, Darvill AG, York WS (2003) Characterization of a tomato protein that inhibits a xyloglucan-specific endoglucanase. Plant J 34:327–338

    Article  CAS  PubMed  Google Scholar 

  • Quiroz-Figueroa FR, Rojas-Herrera R, Galaz-Avalos RM, Loyola-Vargas VM (2006) Embryo production through somatic embryogenesis can be used to study cell differentiation in plants. Plant Cell Tissue Organ Cult 86:285–301

    Article  Google Scholar 

  • Raghavan V (2004) Role of 2,4-dichlorophenoxyacetic acid (2,4-D) in somatic embryogenesis on cultured zygotic embryos of Arabidopsis: cell expansion, cell cycling, and morphogenesis during continuous exposure of embryos to 2,4-D. Am J Bot 91:1743–1756

    Article  CAS  PubMed  Google Scholar 

  • Rakleova G, Keightley A, Panchev I, Tsacheva I, Tchorbadjieva M (2010) Cysteine proteinases and somatic embryogenesis in suspension cultures of orchardgrass (Dactylis glomerata L.) General and Applied Plant Physiol 36:100–109

    CAS  Google Scholar 

  • Rao SR, Ravishankar GA (2002) Plant cell cultures: chemical factories of secondary metabolites. Biotechnol Adv 20:101–153

    Article  CAS  PubMed  Google Scholar 

  • Ribas AF, Dechamp E, Champion A, Bertrand B, Combes MC, Verdeil JL, Lapeypre F, Lashermes P, Etienne H (2011) Agrobacterium-mediated genetic transformation of Coffea arabica (L.) is greatly enhanced by using established embryogenic callus cultures. BMC Plant Biol 11:92. https://doi.org/10.1186/1471-2229-11-92

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sané D, Aberlenc-Bertossi F, Gassama-Dia YK, Sagna M, Trouslot MF, Duval Y, Borgel A (2006) Histocytological analysis of callogenesis and somatic embryogenesis from cell suspensions of date palm (Phoenix dactylifera). Ann Bot 98:301–308

    Article  PubMed  PubMed Central  Google Scholar 

  • Sulová Z, Baran R, Farkaš V (2001) Release of complexed xyloglucan endotransglycosylase (XET) from plant cell walls by transglycosylation reaction with xyloglucan-derived oligosaccharides. Plant Physiol Biochem 39:927–932

    Article  Google Scholar 

  • Takáč T, Pechan T, Šamaj J (2011) Differential proteomics of plant development. J Proteome 74:577–588

    Article  Google Scholar 

  • Tian L, Zhang L, Zhang J, Song Y, Guo Y (2009) Differential proteomic analysis of soluble extracellular proteins reveals the cysteine protease and cystatin involved in suspension-cultured cell proliferation in rice. Biochim Biophys Acta 1794:459–467

    Article  CAS  PubMed  Google Scholar 

  • van der Hoorn RL (2008) Plant proteases: from phenotypes to molecular mechanism. Ann Rev Plant Biol 59:191–223

    Article  Google Scholar 

  • van Hengel AJ, Tadesse Z, Immerzeel P, Schols H, van Kammen A, de Vries SC (2001) N-Acetylglucosamine and glucosamine-containing arabinogalactan proteins control somatic embryogenesis. Plant Physiol 125:1880–1890

    Article  PubMed  PubMed Central  Google Scholar 

  • Vargas TE, García ED, Oropeza M (2005) Somatic embryogenesis in Solanum tuberosum from cell suspension cultures: histological analysis and extracellular protein patterns. J Plant Physiol 162:449–456

    Article  CAS  PubMed  Google Scholar 

  • Vieitez FJ, Ballester A, Vieitez AM (1992) Somatic embryogenesis and plantlet regeneration from cell suspension cultures from Fagus sylvatica L. Plant Cell Rep 11:609–613

    Article  CAS  PubMed  Google Scholar 

  • Wang CX, Wang L, Thomas CR (2004) Modelling the mechanical properties of single suspension-cultured tomato cells. Ann Bot 93:443–453

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yang J, Bi HP, Fan WJ, Zhang M, Wang HX, Zhang P (2011) Efficient embryogenic suspension culturing and rapid transformation of a range of elite genotypes of sweet potato (Ipomoea batatas [L.] Lam). Plant Sci 181:701–711

    Article  CAS  PubMed  Google Scholar 

  • Yang X, Zhang X (2010) Regulation of somatic embryogenesis in higher plants. CRC CR Rev Plant Sci 29:36–57

    Article  CAS  Google Scholar 

  • Yin L, Lan Y, Zhu L (2008) Analysis of the protein expression profiling during rice callus differentiation under different plant hormone conditions. Plant Mol Biol 68:597–617

    Article  CAS  PubMed  Google Scholar 

  • Zhang J, Mab H, Chen S, Ji M, Perl A, Kovacs L, Chen S (2009) Stress response proteins’ differential expression in embryogenic and non-embryogenic callus of Vitis vinifera L. cv. Cabernet Sauvignon—a proteomic approach. Plant Sci 177:103–113

    Article  CAS  Google Scholar 

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Funding

This work was supported by the Centre for Functional Ecology funding and a post-doctoral research fellowship (SFRH/BPD/91461/2012) awarded to S.I.C. by the Fundação para a Ciência e Tecnologia (Portugal).

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Authors and Affiliations

  1. Centre for Functional Ecology, Department of Life Sciences, University of Coimbra, Calçada Martim de Freitas, 3000-456, Coimbra, Portugal

    Ana Alves, André Caeiro, Sandra Isabel Correia & Jorge Canhoto

  2. Department of Life Sciences, University of Coimbra, Calçada Martim de Freitas, 3000-456, Coimbra, Portugal

    Sandra Isabel Correia, Paula Veríssimo & Jorge Canhoto

  3. Centre for Neuroscience and Cell Biology, Faculty of Medicine, University of Coimbra, Rua Larga, 3000-504, Coimbra, Portugal

    Paula Veríssimo

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  1. Ana Alves
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  2. André Caeiro
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  3. Sandra Isabel Correia
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  4. Paula Veríssimo
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  5. Jorge Canhoto
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Correspondence to Sandra Isabel Correia.

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Editor: Rakhi Chaturvedi

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Figure S1

Gelatin zymograph of extracellular proteins from S. betaceum embryogenic (EC1) and non-embryogenic (NEC) suspension cultures; proteolytic activity appears as unstained bands. Molecular weight scale in kilodaltons. (PDF 161 kb)

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Alves, A., Caeiro, A., Correia, S.I. et al. Establishment and biochemical characterization of tamarillo (Solanum betaceum Cav.) embryogenic cell suspension cultures. In Vitro Cell.Dev.Biol.-Plant 53, 606–618 (2017). https://doi.org/10.1007/s11627-017-9864-z

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