Pathogenic Bacteria Target Plant Plasmodesmata to Colonize and Invade Surrounding Tissues.

A hallmark of multicellular organisms is their ability to maintain physiological homeostasis by communicating among cells, tissues, and organs. In plants, intercellular communication is largely dependent on plasmodesmata (PD), which are membrane-lined channels connecting adjacent plant cells. Upon immune stimulation, plants close PD as part of their immune responses. Here, we show that the bacterial pathogen Pseudomonas syringae deploys an effector protein, HopO1-1, that modulates PD function. HopO1-1 is required for P. syringae to spread locally to neighboring tissues during infection. Expression of HopO1-1 in Arabidopsis (Arabidopsis thaliana) increases the distance of PD-dependent molecular flux between neighboring plant cells. Being a putative ribosyltransferase, the catalytic activity of HopO1-1 is required for regulation of PD. HopO1-1 physically interacts with and destabilizes the plant PD-located protein PDLP7 and possibly PDLP5. Both PDLPs are involved in bacterial immunity. Our findings reveal that a pathogenic bacterium utilizes an effector to manipulate PD-mediated host intercellular communication for maximizing the spread of bacterial infection.

Sieve Elements: The Favourite Habitat of Phytoplasmas.

The sieve elements are the only plant compartments, where phytoplasmas can survive and propagate. Therefore, this chapter is focussed on the specific molecular and cell-biological properties of the sieve element. Sieve element-companion cell complexes arise from (pro)cambial mother cells induced by key genes known to be decisive for sieve-element differentiation. The special anatomy, cell biology, and plasma-membrane outfit of sieve elements allows them to act collectively as a tube system that is able to drive a mass flow against the flow induced by transpiration. Plasmodesmal corridors are vital for the translocation of photoassimilates and systemic signals and for survival of the enucleate sieve elements. Of paramount importance is the Ca2+-dependent gating of plasmodesmata by callose and proteins. Hence, some of the complex, regulatory mechanisms to maintain Ca2+ homoeostasis in sieve elements are presented. Finally, the peculiarities of the chemical and physical sieve-element environment offered to phytoplasmas are discussed.

The Fusarium oxysporum Avr2-Six5 Effector Pair Alters Plasmodesmatal Exclusion Selectivity to Facilitate Cell-to-Cell Movement of Avr2.

Pathogens use effector proteins to manipulate their hosts. During infection of tomato, the fungus Fusarium oxysporum secretes the effectors Avr2 and Six5. Whereas Avr2 suffices to trigger I-2-mediated cell death in heterologous systems, both effectors are required for I-2-mediated disease resistance in tomato. How Six5 participates in triggering resistance is unknown. Using bimolecular fluorescence complementation assays we found that Avr2 and Six5 interact at plasmodesmata. Single-cell transformation revealed that a 2xRFP marker protein and Avr2-GFP only move to neighboring cells in the presence of Six5. Six5 alone does not alter plasmodesmatal transduction as 2xRFP was only translocated in the presence of both effectors. In SIX5-expressing transgenic plants, the distribution of virally expressed Avr2-GFP, and subsequent onset of I-2-mediated cell death, differed from that in wild-type tomato. Taken together, our data show that in the presence of Six5, Avr2 moves from cell to cell, which in susceptible plants contributes to virulence, but in I-2 containing plants induces resistance.

Histochemical Analyses Reveal That Stronger Intrinsic Defenses in Gossypium barbadense Than in G. hirsutum Are Associated With Resistance to Verticillium dahliae.

Verticillium wilt, caused by Verticillium dahliae Kleb., is a serious threat to cotton (Gossypium spp.) crop production. To enhance our understanding of the plant's complex defensive mechanism, we examined colonization patterns and interactions between V. dahliae and two cotton species, the resistant G. barbadense and the susceptible G. hirsutum. Microscopic examinations and grafting experiments showed that the progression of infection was restricted within G. barbadense. At all pre- and postinoculation sampling times, levels of salicylic acid (SA) were also higher in that species than in G. hirsutum. Comparative RNA-Seq analyses indicated that infection induced dramatic changes in the expression of thousands of genes in G. hirsutum, whereas those changes were fewer and weaker in G. barbadense. Investigations of the morphological and biochemical nature of cell-wall barriers demonstrated that depositions of lignin, phenolic compounds, and callose were significantly higher in G. barbadense. To determine the contribution of a known resistance gene to these processes, we silenced GbEDS1 and found that the transformed plants had decreased SA production, which led to the upregulation of PLASMODESMATA-LOCATED PROTEIN (PDLP) 1 and PDLP6. This was followed by a decline in callose deposition in the plasmodesmata, which then led to increased pathogen susceptibility. This comparison between resistant and susceptible species indicated that both physical and chemical mechanisms play important roles in the defenses of cotton against V. dahliae.

Investigating plasmodesmata genetics with virus-induced gene silencing and an agrobacterium-mediated GFP movement assay.

Plasmodesmata (PD) are channels that connect the cytoplasm of adjacent plant cells, permitting intercellular transport and communication. PD function and formation are essential to plant growth and development, but we still know very little about the genetic pathways regulating PD transport. Here, we present a method for assaying changes in the rate of PD transport following genetic manipulation. Gene expression in leaves is modified by virus-induced gene silencing. Seven to ten days after infection with Tobacco rattle virus carrying a silencing trigger, the gene(s) of interest is silenced in newly arising leaves. In these new leaves, individual cells are then transformed with Agrobacterium to express GFP, and the rate of GFP diffusion via PD is measured. By measuring GFP diffusion both within the epidermis and between the epidermis and mesophyll, the assay can be used to study the effects of silencing a gene(s) on PD transport in general, or transport through secondary PD specifically. Plant biologists working in several fields will find this assay useful, since PD transport impacts plant physiology, development, and defense.

Airborne signals from a wounded leaf facilitate viral spreading and induce antibacterial resistance in neighboring plants.

Many plants release airborne volatile compounds in response to wounding due to pathogenic assault. These compounds serve as plant defenses and are involved in plant signaling. Here, we study the effects of pectin methylesterase (PME)-generated methanol release from wounded plants ("emitters") on the defensive reactions of neighboring "receiver" plants. Plant leaf wounding resulted in the synthesis of PME and a spike in methanol released into the air. Gaseous methanol or vapors from wounded PME-transgenic plants induced resistance to the bacterial pathogen Ralstonia solanacearum in the leaves of non-wounded neighboring "receiver" plants. In experiments with different volatile organic compounds, gaseous methanol was the only airborne factor that could induce antibacterial resistance in neighboring plants. In an effort to understand the mechanisms by which methanol stimulates the antibacterial resistance of "receiver" plants, we constructed forward and reverse suppression subtractive hybridization cDNA libraries from Nicotiana benthamiana plants exposed to methanol. We identified multiple methanol-inducible genes (MIGs), most of which are involved in defense or cell-to-cell trafficking. We then isolated the most affected genes for further analysis: ß-1,3-glucanase (BG), a previously unidentified gene (MIG-21), and non-cell-autonomous pathway protein (NCAPP). Experiments with Tobacco mosaic virus (TMV) and a vector encoding two tandem copies of green fluorescent protein as a tracer of cell-to-cell movement showed the increased gating capacity of plasmodesmata in the presence of BG, MIG-21, and NCAPP. The increased gating capacity is accompanied by enhanced TMV reproduction in the "receivers". Overall, our data indicate that methanol emitted by a wounded plant acts as a signal that enhances antibacterial resistance and facilitates viral spread in neighboring plants.