Introduction

Hundreds of millions of years of co-evolution between plants and fungi have resulted in over 99% of orchids using partial mycoheterotrophy in which young plants obtain carbon nutrients from fungi prior to the development of green leaves (Yuan et al. 2018). Gastrodia in Orchidaceae represents an unusual fully mycoheterotrophic genus (Leake 1994). For efficient cultivation, G. elata first requires a symbiotic association with Mycena spp. for seed germination and later with Armillaria for tuber formation and sexual reproduction (Kim et al. 2006; Lan, Xu & Li 1994; Xu & Fan 2001). Without photosynthetic ability, G.elata requires carbon fromArmillaria fungus and degraded hyphae to complete its ~36-month life cycle (Zhang & Li 1980). This unique symbiotic interaction of G. elata provides a valuable system to explore how carbon is distributed and allocated in mycoheterotrophic plants.
In symbiotic autotrophic plants, high carbon transfer has been documented in the interface between host roots and the arbuscular mycorrhizae (AM) fungal partner (Vandenkoornhuyse et al. 2007). The morphology is similar to Armillaria during mycoheterotrophic symbiosis. The carbon source has been shown to be key to establish optimal symbiosis (Hennion et al. 2019; Kiers et al. 2011). In particular, soluble sugars, such as sucrose or hexoses, are probably the major form of carbon flow (Garcia, Doidy, Zimmermann, Wipf & Courty 2016; Gutjahr, Novero, Welham, Wang & Bonfante 2011). Sugar transporters have been identified that localize on adjacent plant and fungal cells and these may fine-tune sugar exchange ensuring benefits for both partners in this symbiotic relationship (Doidy et al. 2012a; Hennion et al.2019). These transport systems include gene families encoding SUT/SUCs (sucrose transporters), MSTs (monosaccharide transporters) and SWEETs (sugars will eventually be exported transporter) (Doidy et al.2012a; Garcia et al. 2016; Manck-Götzenberger & Requena
2016). SUT transporters represent the most likely candidate to mediate sucrose transport during plant-fungal symbiosis due to increased expression of the SUT genes upon symbiosis (Boldt et al. 2011; Doidy et al. 2012b).
SUTs, encoded by a small gene family, are classified into four major clades (SUT1, SUT2-IIA, SUT2-IIB, SUT4) and mostly function as sucrose/H+ symporters (Doidy et al. 2012b; Julius, Leach, Tran, Mertz & Braun 2017; Kühn & Grof 2010). The well- characterized clade 1 includes Arabidopsis At SUC2 (Gottwald, Krysan, Young, Evert
& Sussman 2000; Srivastava, Ganesan, Ismail & Ayre 2008) and potatoSt SUT1 (Lemoine, Kühn, Thiele, Delrot & Frommer 1996), the key sucrose importer that mediates phloem loading in source leaves for long-distance sugar transport (Kühn et al. 2003). In contrast, carriers in clades 2 and 4 mobilize sucrose in sink organs for their growth. For example, OsSUT1 and HvSUT1 (Furbank et al. 2001; Weschke et al. 2000), members of clade 2, are expressed in developing grains to transport sucrose into the endosperm for grain filling (Radchuk et al. 2017; Scofield et al.2002). Potato StSUT4 , a clade 4 member, mediates sucrose uptake across the plasma membrane to initiate tuberization and flowering (Chincinska et al. 2008; Weise et al. 2000).
SUT transporters also participate in sucrose allocation between autotrophic plants and symbiotic fungi. Expression of several SUT genes is greatly induced upon inoculation of Medicago and tomato roots with AM fungi (Boldt et al. 2011; Doidy et al. , 2012b). Overexpression of the potato St SUT1 increased AM colonization under high available phosphate, likely due to increased sugar allocation to roots for export to AM fungi (Gabriel-Neumann, Neumann, Leggewie & George 2011). Similarly, increased sucrose export by silencing expression of a periarbuscular membrane retriever, Sl SUT2, promoted mycorrhization in tomato roots (Bitterlich, Krügel, Boldt- Burisch, Franken & Kühn 2014). These studies illustrate the importance of sucrose
allocation and the activity of SUT sucrose transporters for optimal plant-fungus symbiosis. A similar sucrose distribution system may also operate in mycoheterotrophic plant cells and require a SUT transport mechanism.
To examine the carbon flow mechanism in mycoheterotrophic plants, we utilized the unique Gastrodia-Armillaria symbiotic system to identify two SUT sucrose transporters, GeSUT4 and GeSUT3 , which were highly expressed in G. elata young symbiotic tubers. Transport analysis and localization experiments revealed thatGe SUT4 functions as a sucrose-specific importer on the plasma membrane and the tonoplast in vivo . Based on functional characterizations in Arabidopsis, we propose that Ge SUT4 participates in fungal derived sucrose import during mycoheterotrophic tuber formation. A model of this process is proposed.