GeSUT4 is a sucrose-specific active
importer in G.
elata
We
also identified genes encoding sucrose transporters, Ge SUT4 andGe SUT3, that are expressed in mycoheterotrophic Gastradiacells (Fig. 3a, b). The high expression of GeSUT genes in young
and elongating tubers (stages 2A-2B and 4) coincides with high levels of
sucrose, but not of glucose, in developing tubers (Fig. 1). As been
observed in most plant-fungal interfaces (Doidy et al. 2012a;
Garcia et al. 2016; Hennion et al. 2019), distinct
expression of GeSUT genes occurred in the absence of any
symplastic connections in infected cortical cells and large cells (Fig.
2 and Fig. 5c). The sucrose- specific transport activity ofGe SUT4 was demonstrated by complementation and uptake kinetics in
the yeast mutant (Fig. 4). The sensitivity of Ge SUT4-mediated
sucrose import to alkaline pH and the protonophore CCCP confirmed that
this transporter is a proton-dependent transporter (Fig. 4e), as shown
for most characterized SUT4 type carriers (Kühn & Grof 2010; Reinders,
Sivitz & Ward 2012; Schulz et al. 2011; Weise et al.2000). Considering the high amino acid identity (up to 58 %) to theOs SUT2 symporter (Eom et al. 2011), it is possible thatGe SUT4 functions as a proton-symporter. The presence of an active
uptake mechanism like Ge SUT4 can also be supported by expression
of various P-type ATPases in the G. elata genome (Yuan et
al. 2018). The absence of Ge SUT3 transport activity in yeast may
be due to inappropriate expression in a heterologous system andGe SUT3 may still be involved in sucrose homeostasis in
vivo .
GeSUT4
exhibits a high substrate affinity (KM ~2.5 mM) for
sucrose (Fig. 4c), similar to its close homolog in rice, OsSUT2
(KM ~1.8mM) (Eom et al. 2011). The feature nicely
fits with the low sucrose contents measured in the plant-microbe
interfaces (Naher, Radziah, Halimi, Shamsuddin & Razi 2008). Sucrose
transporters characterized in some pathogenic fungal cells also exhibit
high affinity to sucrose (Sutton, Henry & Hall 1999; Wahl, Wippel,
Goos, Kamper & Sauer 2010). The transport properties of GeSUT4
would enable strong competition for carbon nutrients from the symbiotic
fungal cells and could provide a strategy to avoid rapid metabolism of
sucrose into hexoses during carbon allocation.