All multi-cellular organisms require effective intercellular communication to coordinate cellular processes. In plants, cell-to-cell contact is restricted by the presence of a rigid cell wall.
Nevertheless, cells are connected via plasmodesmata (PD), membrane-lined, cell wall channels that provide cytoplasmic continuity and form a tightly regulated system that allows exchange of molecules between neighboring cells. (Lee and Lu, 2011; Maule et al., 2011; Burch-Smith and Zambryski, 2012). In general, PD are lined with PM and a tube of the ER, the so-called desmotuble, that is tightly coiled by reticulons, runs across the pore (Figure 5). Callose, a plant specific polysaccharide consisting of β-1,3 linked glucose, may be deposited in the neck regions
of the tunnel to restrict the PD-flux (Maule et al., 2011; Burch-Smith and Zambryski, 2012;
Maule et al., 2012). The cytoplasmic sleeve, the space between PM and desmotubule is filled with cytoskeletal proteins that are important for the PD structure and form and modulate the size exclusion limit (SEL) for transport through the PD (Christensen et al., 2009; Chen et al., 2010).
In addition to the cytoplasmic sleeve, the membrane of the desmotubule as well as its lumen serve as trafficking pathways (Guenoune-Gelbart et al., 2008; Barton et al., 2011). Recent analyses of PD-enriched cell wall fractions suggest that PD-associated membranes have distinct features. The studies revealed the presence of proteins like remorins (Raffaele et al., 2009), tetraspanins (Salmon and Bayer, 2012), RLKs and GPI-anchored proteins (Fernandez-Calvino et al., 2011) in PD. Interestingly, PD membranes are enrichment in sterols and sphingolipids with very long chain saturated fatty acids (Grison et al., 2015). This lipid profile is reminiscent of detergent-insoluble PM microdomains which have been found to typically harbor RLKs and GPI-anchored proteins (Thomas et al., 2008; Raffaele et al., 2009; Simpson et al., 2009) (Figure 5). The microdomain-like nature of PD membranes may be important for its function, such as in sorting and recruiting associated proteins.
The structure of the PM allows small uncharged molecules to diffuse through. Various proteins, including ion channels, protein pumps and carrier proteins help large or charged molecules pass through the cell membrane. Transport through PD is presumed to be passive but there is evidence that they facilitate the active transposition of so called non-cell autonomous proteins (NCAPs) acting in developmental processes (Haywood et al., 2002). Moreover, developmental control includes hormone signaling, transcription factor and sRNA/mRNA trafficking between cells and tissues (Zambryski and Crawford, 2000).
PD are involved in several processes one of which is the regulation of plant growth and development. They also may help to determine a program of cell differentiation, such as sealing off root and stem epidermal cells from the rest of the plant (Zambryski and Crawford, 2000;
Burch-Smith et al., 2011; Burch-Smith and Zambryski, 2012). Moreover, by regulating their diameter PD play an important role to establish and maintain physiological gradients between cells. The translocation of molecules is limited by the SEL (Xu and Jackson, 2010; Xu et al., 2012). Molecules smaller than the SEL of plasmodesmata are able to move freely through the cytoplasmic channel of plasmodesmata by simple diffusion. The SEL can be modified due to environmental changes such as cytoplasmic calcium levels or in response to changes in turgor pressure between cells (Burch-Smith et al., 2011; Burch-Smith and Zambryski, 2012). The signaling processes are tightly regulated by limiting the active and passive transport through
Introduction
PD. Interestingly, also auxin could be linked to locally down regulated symplastic permeability by inducing callose deposition at PD (Han et al., 2014). Recent research identified PD-resident proteins involved in callose homeostasis that are associated with the regulation of the PD-flux in both developmental and disease-related contexts (Guseman et al., 2010; Vaten et al., 2011;
Maule et al., 2012).
Figure 5: Simplified model of a plasmodesma.
The illustration shows the structural domains of the PD pore. The cellulosic or pectin rich cell wall, the PM and the desmotubule are depicted. Different membrane domains are found at PD (A) the PM, (B) the desmotuble coiled by reticulons, (C) remorin enriched microdomains with GPI-anchored proteins or (E) receptor-like proteins and (D) tetraspanin-enriched microdomains that provide a platform for receptor function. Figure adapted from Maule et al.
(2011).
1.5.1 The function of plasmodesmata in plant innate immunity
A number of pathogens move through plasmodesmata to colonize plant tissues. For expample, most plant viruses use their movement proteins to modify PD and spread from cell to cell (Ueki and Citovsky, 2011; Tilsner et al., 2013). Other pathogens like the hemibiotrophic fungus Magnaporthe oryzae, exploit these structures by growing through PD to infect the neighboring cells (Kankanala et al., 2007). Strategies to recognize and remodel PD by pathogens allow rapid entry into neighboring cells by keeping the PM intact and thereby prevent plant defense.
Therefore PD are ideal locations for structural components of plants innate immunity. Indeed, PD are membrane domains rich in receptor proteins (Fernandez-Calvino et al., 2011). Guarding the PD tunnel with several types of receptor proteins is a plant strategy to counteract pathogens using PD as a route of cell-to-cell movement. Arabidopsis contains eight PD-Located Proteins (PDLPs), a family of PD-specific RLPs with cysteine-rich ectodomains (Thomas et al., 2008; Lee et al., 2011). Overexpression of PDLP5 causes callose deposition at PD and consequently decreased PD transport. It also causes over-accumulation of SA and an associated cell death phenotype. Moreover PDLP5 overexpression restricts proliferation of Pseudomonas syringae and tobacco mosaic virus (TMV), presumably reduced PD connectivity and increased SA levels (Lee et al., 2011). The chitin-binding LysM-RLP LYM2 was also found in a proteomic study on PD proteins (Fernandez-Calvino et al., 2011). Analysis of plants expressing mCitrine-LYM2 fusion proteins indicated that LYM2 is distributed throughout the PM, but shows areas of higher accumulation at PD (Faulkner et al., 2013). Chitin treatment leads to a reduction in PD connectivity in wild type Arabidopsis plants. Interestingly, lym2 mutants were no longer able to restrict transport through PD upon chitin treatment, whereas this response was normal in cerk1-2. These results suggest a CERK1-independent role of LYM2 in PD regulation, which is also important for resistance to fungal pathogens (Faulkner et al., 2013). The fact that the PD-located proteins PDLP5 and LYM2 confer resistance to plant pathogens emphasize the crucial role of PD in plant defense (Lee and Lu, 2011).
Introduction