Introduction
Terpenoids constitute the largest and most diverse class of chemical
substances that play major roles in plant primary and secondary
metabolisms (Gershenzon & Dudareva, 2007; Nagel, Berasategui, Paetz,
Gershenzon & Schmidt, 2014; Trapp & Croteau, 2001). Two spatially
separated pathways exist for the biosynthesis of terpenoids; the
mevalonate (MVA) pathway takes place in cytosol and peroxisomes, whereas
the methylerythritol phosphate (MEP) pathway is active in plastids
(Dudareva, Klempien, Muhlemann, & Kaplan, 2013; Nagegowda, 2010). Both
pathways form the C5 compounds isopentenyl diphosphate (IPP) and its
allylic isomer, dimethylallyl diphosphate (DMAPP), which act as
precursors for the biosynthesis of all terpenoids (McGarvey & Croteau,
1995). Whereas the MVA pathway produces sesquiterpenes (C15), irregular
terpenes and geranyllinalool, the MEP pathway forms hemiterpenes (C5),
monoterpenes (C10), diterpenes (C20) and volatile carotenoid derivatives
(McGarvey & Croteau, 1995; Muhlemann, Klempien & Dudareva, 2014).
Terpenoids can be stored in specialized structures such as resin ducts
in many conifers (Gershenzon & Croteau, 1991; Wu & Hu, 1997) or
directly emitted into the environment (Loreto et al., 2001). In storing
plants, terpenoids usually make up 1-2% of plant dry weight, in some
special cases shares of up to 15-20% were observed (Blanch, Penuelas,
Sardans & Llusia, 2009). Terpenoids exert key ecological functions for
plants, such as defence against predators, pathogens or competitors,
inter- and intraspecific communication as well as protection against
abiotic stress (Gershenzon & Dudareva, 2007; Loreto & Schnitzler,
2010; Loreto, Pollastri, Fineschi & Velikova, 2014; Martin, Gershenzon
& Bohlmann, 2003; Vickers et al., 2009).
According to present knowledge, there are two sources driving terpenoid
emission from leaves of plants. (i) De novo biosynthesis controls
emission of isoprene and terpenoids from non-storing plants such asQuercus ilex . Production of these compounds is tightly coupled to
photosynthesis and therefore shows similar dependencies on environmental
parameters such as light availability and temperature (Ghirardo et al.,
2010; Loreto et al., 1996; Loreto, Nascetti, Graverini & Mannozzi,
2000; Tingey, Manning, Grothaus & Burns, 1979). On the other hand, (ii)
the release of leaf-internally stored terpenoids is purely temperature
dependent because temperature controls the corresponding saturation
vapour pressures of the emitted compounds (Ghirardo et al., 2010;
Guenther, Zimmerman, Harley, Monson & Fall, 1993; Wu et al., 2017).
As a typical conifer, Norway spruce (Picea abies ) stores large
amounts of terpenoids in resin ducts of various tissues, which are at
least partially produced in the epithelial cells of the resin ducts and
mesophyll tissue (Schürmann, Ziegler, Kotzias, Schönwitz &
Steinbrecher, 1993). 13C-labelling approaches
demonstrated that about one third of the emitted monoterpenoids in
spruce are derived from de novo biosynthesis whereas the
remainder is released from storage pools in a temperature depended
manner (Ghirardo et al., 2010; Grabmer et al., 2006). Recent studies
provided indirect hints that besides de novo biosynthesis and/or
release from storage pools, xylem-transported terpenoids might
contribute to emission from Norway spruce needles. Bourtsoukidis et al.
(2012, 2014) observed that sesquiterpene (SQT) emission from Norway
spruce strongly correlated with relative humidity. At high relative
humidity - and consequently reduced transpiration rates - SQT emission
was slowed down and vice versa when relative humidity was below
50% (Bourtsoukidis et al., 2012). In agreement with this finding,
Filella, Wilkinson, Llusia, Hewitt & Peñuelas (2007) demonstrated that
the emissions of most volatile organic compounds including monoterpenes
from Norway spruce were well related to stomatal conductance and
transpiration. This effect may in part be explained by a contribution of
the tree’s sap flow to terpenoid emissions. Moreover, Bäck et al. (2012)
found the largest SQT pools in Pinus sylvestris not in needles
but below the bark, where SQTs constitute a toxic barrier for bark
beetles and function as stress defense agents. Such terpenes might
partially be released into the xylem sap. This assumption is in
consistence with observations of Kuroda (1991) who observed a strong
link between xylem cavitation in pine wilt disease infected Pinustrees and terpene abundance in the xylem. The author hypothesizes that
cavitation is caused by higher terpene content in the xylem sap and the
thereby diminished adhesion forces. A strong induction of terpene
biosynthesis in the developing xylem by different stresses has also been
demonstrated in Norway spruce (Martin, Tholl, Gershenzon & Bohlmann,
2002).
In this study, we aim at clarifying whether or not terpenoids are
transported in the xylem sap of Norway spruce. Moreover, we aimed to
obtain information on possible sources of xylem-transported terpenoids.
In addition, we tried to find hints if xylem-transported terpenoids
might contribute to terpenoid emission from Norway spruce
needles.Therefore, we analyzed terpenoid emission rates as well as
terpenoid contents and compositions in root, bark and wood of the same
Norway spruce trees for comparison with xylem sap terpenoids.