Arabinogalactan-proteins stimulate somatic embryogenesis and plant propagation of Pelargonium sidoides
Introduction
Arabinogalactan-proteins are a group of hydroxyproline-rich proteoglycans present in the extracellular matrix of plants, sometimes linked to the plasma membrane by a glycosylphosphatidylinositol anchor. They have been isolated from plant cell walls, mucilages and gums as well as from cell cultures of various plants. The relatively small protein core of AGPs (normally around 10% of the molecule) is covalently linked to arabinogalactans which have a typical structure consisting mainly of 3-,6- and 3,6-linked β-d-Galp residues, substituted with α-l-Araf residues (for reviews see Ellis, Egelund, Schultz, & Bacic, 2010; Nothnagel, 1997). Different studies indicate a range of functions of AGPs associated with growth and development, including cell division, elongation and proliferation, xylem differentiation, programmed cell death, plant microbe interactions, and also embryogenesis (Ellis et al., 2010, Seifert and Roberts, 2007).
The medicinal plant Pelargonium sidoides is native to South Africa and its roots are used for the production of herbal medicinal products for the treatment of acute bronchitis (for review see Brendler & van Wyk, 2008). Recently, it has been demonstrated, that an extract from roots of P. sidoides stimulates the innate immune defense by activation of human monocytes (Witte, Koch, Volk, Wolk, & Sabat, 2015). Until today, the plant material partly originates from wild collections. With regard to species protection in combination with an increasing demand of plant material for the industrial production, we developed a new method for plant reproduction of P. sidoides by direct somatic embryogenesis (SE; Duchow, Blaschek, & Classen, 2015).
SE has been described for different species as good method for in vitro multiplication and conservation of plants (Pereira-Netto et al., 2007; Steinmacher, Guerra, Saare-Surminski, & Lieberei, 2011). Based on the knowledge, that AGPs stimulate somatic embryogenesis in different species (Pereira-Netto et al., 2007; Poon, Heath, & Clarke, 2012; Simonović et al., 2015, Van Hengel et al., 2001), we isolated and characterized AGPs from roots of the native plants as well as from callus and suspension cultures generated from roots and shoots of germinating seeds of P. sidoides and investigated their potential to stimulate SE and thereby in vitro plant reproduction and plant regeneration of this important medicinal plant.
Section snippets
Material
Pelargonium sidoides DC (Geraniaceae) has been grown in the Pharmaceutical Garden of the Pharmaceutical Institute of the University of Kiel, Germany. Callus cultures from shoots and roots of germinating seeds as well as suspension cultures derived from calli were established as described by Duchow, Blaschek, & Classen (2014).
AGPs from roots
AGPs were isolated from powdered roots (100 g) by pre-extraction with acetone 60% (v/v) at 4 °C. After rinsing with acetone 60% for 3 times, the suspension was filtered and
Yield of AGPs
Yield of AGPs from cell cultures after precipitation with Yariv́s reagent varied between 0.4 and 0.9% in relation to freeze-dried cells and was higher for callus culture (0.9%) compared to suspension culture (in mean 0.44%, Table 1). This is in a range comparable to AGP yields from other plant cell cultures. In suspension culture medium of cultures of Echinacea purpurea, AGPs accounted for 0.43-0.80% of dry weight of the cell material (Classen, 2007). Yields from cells and medium of suspension
Conclusion
Arabinogalactan-proteins from roots and cell cultures have been structurally characterized and shown to have potential to enhance plant propagation of the medicinal plant P. sidoides by stimulation of somatic embryogenesis.
Acknowledgments
The authors thank F. Geerds and C. Ramisch for assistance in cell culture and somatic embryogenesis as well as S. Offt and O. Zoelck for care of plants in the greenhouse.
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