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Specific targeting of a plasmodesmal protein affecting cell-to-cell communication.

Thomas CL, Bayer EM, Ritzenthaler C, Fernandez-Calvino L, Maule AJ - PLoS Biol. (2008)

Bottom Line: We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking.These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication.They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.

View Article: PubMed Central - PubMed

Affiliation: John Innes Centre, Norwich Research Park, Colney, Norwich, United Kingdom.

ABSTRACT
Plasmodesmata provide the cytoplasmic conduits for cell-to-cell communication throughout plant tissues and participate in a diverse set of non-cell-autonomous functions. Despite their central role in growth and development and defence, resolving their modus operandi remains a major challenge in plant biology. Features of protein sequences and/or structure that determine protein targeting to plasmodesmata were previously unknown. We identify here a novel family of plasmodesmata-located proteins (called PDLP1) whose members have the features of type I membrane receptor-like proteins. We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking. PDLP1a is targeted to plasmodesmata via the secretory pathway in a Brefeldin A-sensitive and COPII-dependent manner, and resides at plasmodesmata with its C-terminus in the cytoplasmic domain and its N-terminus in the apoplast. Using a deletion analysis, we show that the single transmembrane domain (TMD) of PDLP1a contains all the information necessary for intracellular targeting of this type I membrane protein to plasmodesmata, such that the TMD can be used to target heterologous proteins to this location. These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication. They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.

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PDLP1a Targeting to Plasmodesmata(A) shows a cartoon of a plasmodesma. The plasmodesma is seen as a PM-lined channel through the cell wall containing a central ER-derived rod-shaped desmotubule (DT). The location of possible proteinaceous components (arrows), as visualised by electron microscopy [6], in the symplastic channel is illustrated.(B–D) show subcellular location of PDLP1a:GFP following transgenic expression in Arabidopsis. Following expression of PDLP1a:GFP under the control of the CaMV 35S promoter (B) or the PDLP1a native promoter (C), optical sections through the lower epidermis show the targeting of PDLP1a to plasmodesmata. The upper images show the typical punctate distribution of plasmodesmata along the cell wall of adjoining cells. The lower images show an overlay of the fluorescence on the bright field image of the same area to identify the location of the cell walls.(D and E) PDLP1a:GFP remained at punctate structures within the cell wall after plasmolysis, as shown for epidermal cells (D), and on the adjoining wall between adjacent spongy mesophyll cells (E). An overlay of the fluorescence on the bright field image of the same area is shown; asterisks (*) demarcate the retracted protoplast, and arrows identify PDLP1a:GFP retained on the cell wall.Bar indicates 10 μm.
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pbio-0060007-g001: PDLP1a Targeting to Plasmodesmata(A) shows a cartoon of a plasmodesma. The plasmodesma is seen as a PM-lined channel through the cell wall containing a central ER-derived rod-shaped desmotubule (DT). The location of possible proteinaceous components (arrows), as visualised by electron microscopy [6], in the symplastic channel is illustrated.(B–D) show subcellular location of PDLP1a:GFP following transgenic expression in Arabidopsis. Following expression of PDLP1a:GFP under the control of the CaMV 35S promoter (B) or the PDLP1a native promoter (C), optical sections through the lower epidermis show the targeting of PDLP1a to plasmodesmata. The upper images show the typical punctate distribution of plasmodesmata along the cell wall of adjoining cells. The lower images show an overlay of the fluorescence on the bright field image of the same area to identify the location of the cell walls.(D and E) PDLP1a:GFP remained at punctate structures within the cell wall after plasmolysis, as shown for epidermal cells (D), and on the adjoining wall between adjacent spongy mesophyll cells (E). An overlay of the fluorescence on the bright field image of the same area is shown; asterisks (*) demarcate the retracted protoplast, and arrows identify PDLP1a:GFP retained on the cell wall.Bar indicates 10 μm.

Mentions: Plasmodesmata are channels that cross the cell wall and establish symplastic continuity throughout most of the plant. Their importance has been highlighted by the range of diverse non–cell-autonomous functions that depend on intercellular macromolecular communication through plasmodesmata. Hence, a range of transcription factors in the shoot apical meristem and at the root tip have functional roles in cells other than those in which they were produced [1,2]. Similarly, some small RNAs generated as part of the RNA silencing pathway can act non–cell-autonomously [3]. Last, plant virus pathogens, which are restricted to the symplast, must use plasmodesmata to invade neighbouring cells [4], and very recently [5] it has been proposed that biotrophic fungi may exploit plasmodesmata during tissue invasion. All of these macromolecules and organisms are above the experimentally defined normal size-exclusion limits for plasmodesmata, which points to highly regulated plasmodesmal processes for their recognition and transport between cells. This is exemplified by plant viruses, all of which encode so-called movement proteins (MPs) that interact with and modify the properties of these structures to allow the passage of virus particles or other forms of ribonucleoprotein complexes [4]. The organisational complexity of plasmodesmata remains a matter of speculation constrained by the interpretation of electron microscopical images [6]. Briefly, they comprise plasma membrane (PM)-lined channels that cross the cell wall to join adjacent cells symplastically. They contain an axial-appressed membrane element derived from the endoplasmic reticulum (ER), called the desmotubule, and may contain proteinaceous spoke-like structures that cross the cytoplasmic sleeve between the PM and desmotubule (Figure 1A). Despite the crucial role of plasmodesmata in growth and development, plant defence, and pathogenesis, almost nothing is known about the integral components of plasmodesmata or plasmodesmal biogenesis. From the identification of proteins interacting with viral MPs, coimmunolocalisation studies applied to candidate proteins, and proteomics approaches, a number of proteins have been identified in association with plasmodesmata [7]. These include cytoskeletal elements (e.g., actin and myosin VIII), proteins found in the ER (e.g., calreticulin) (refs in [7]), a casein kinase 1 that phosphorylates tobacco mosaic virus (TMV) MP [8], a β-1,3-glucanase [9], and class 1 reversibly glycosylated proteins (C1RGPs) [10]. The lack of identification of the constituent components of plasmodesmata has remained an outstanding challenge in plant biology and has hindered a fuller understanding of the non–cell-autonomous control of plant development and the processes of tissue invasion by pathogens.


Specific targeting of a plasmodesmal protein affecting cell-to-cell communication.

Thomas CL, Bayer EM, Ritzenthaler C, Fernandez-Calvino L, Maule AJ - PLoS Biol. (2008)

PDLP1a Targeting to Plasmodesmata(A) shows a cartoon of a plasmodesma. The plasmodesma is seen as a PM-lined channel through the cell wall containing a central ER-derived rod-shaped desmotubule (DT). The location of possible proteinaceous components (arrows), as visualised by electron microscopy [6], in the symplastic channel is illustrated.(B–D) show subcellular location of PDLP1a:GFP following transgenic expression in Arabidopsis. Following expression of PDLP1a:GFP under the control of the CaMV 35S promoter (B) or the PDLP1a native promoter (C), optical sections through the lower epidermis show the targeting of PDLP1a to plasmodesmata. The upper images show the typical punctate distribution of plasmodesmata along the cell wall of adjoining cells. The lower images show an overlay of the fluorescence on the bright field image of the same area to identify the location of the cell walls.(D and E) PDLP1a:GFP remained at punctate structures within the cell wall after plasmolysis, as shown for epidermal cells (D), and on the adjoining wall between adjacent spongy mesophyll cells (E). An overlay of the fluorescence on the bright field image of the same area is shown; asterisks (*) demarcate the retracted protoplast, and arrows identify PDLP1a:GFP retained on the cell wall.Bar indicates 10 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2211546&req=5

pbio-0060007-g001: PDLP1a Targeting to Plasmodesmata(A) shows a cartoon of a plasmodesma. The plasmodesma is seen as a PM-lined channel through the cell wall containing a central ER-derived rod-shaped desmotubule (DT). The location of possible proteinaceous components (arrows), as visualised by electron microscopy [6], in the symplastic channel is illustrated.(B–D) show subcellular location of PDLP1a:GFP following transgenic expression in Arabidopsis. Following expression of PDLP1a:GFP under the control of the CaMV 35S promoter (B) or the PDLP1a native promoter (C), optical sections through the lower epidermis show the targeting of PDLP1a to plasmodesmata. The upper images show the typical punctate distribution of plasmodesmata along the cell wall of adjoining cells. The lower images show an overlay of the fluorescence on the bright field image of the same area to identify the location of the cell walls.(D and E) PDLP1a:GFP remained at punctate structures within the cell wall after plasmolysis, as shown for epidermal cells (D), and on the adjoining wall between adjacent spongy mesophyll cells (E). An overlay of the fluorescence on the bright field image of the same area is shown; asterisks (*) demarcate the retracted protoplast, and arrows identify PDLP1a:GFP retained on the cell wall.Bar indicates 10 μm.
Mentions: Plasmodesmata are channels that cross the cell wall and establish symplastic continuity throughout most of the plant. Their importance has been highlighted by the range of diverse non–cell-autonomous functions that depend on intercellular macromolecular communication through plasmodesmata. Hence, a range of transcription factors in the shoot apical meristem and at the root tip have functional roles in cells other than those in which they were produced [1,2]. Similarly, some small RNAs generated as part of the RNA silencing pathway can act non–cell-autonomously [3]. Last, plant virus pathogens, which are restricted to the symplast, must use plasmodesmata to invade neighbouring cells [4], and very recently [5] it has been proposed that biotrophic fungi may exploit plasmodesmata during tissue invasion. All of these macromolecules and organisms are above the experimentally defined normal size-exclusion limits for plasmodesmata, which points to highly regulated plasmodesmal processes for their recognition and transport between cells. This is exemplified by plant viruses, all of which encode so-called movement proteins (MPs) that interact with and modify the properties of these structures to allow the passage of virus particles or other forms of ribonucleoprotein complexes [4]. The organisational complexity of plasmodesmata remains a matter of speculation constrained by the interpretation of electron microscopical images [6]. Briefly, they comprise plasma membrane (PM)-lined channels that cross the cell wall to join adjacent cells symplastically. They contain an axial-appressed membrane element derived from the endoplasmic reticulum (ER), called the desmotubule, and may contain proteinaceous spoke-like structures that cross the cytoplasmic sleeve between the PM and desmotubule (Figure 1A). Despite the crucial role of plasmodesmata in growth and development, plant defence, and pathogenesis, almost nothing is known about the integral components of plasmodesmata or plasmodesmal biogenesis. From the identification of proteins interacting with viral MPs, coimmunolocalisation studies applied to candidate proteins, and proteomics approaches, a number of proteins have been identified in association with plasmodesmata [7]. These include cytoskeletal elements (e.g., actin and myosin VIII), proteins found in the ER (e.g., calreticulin) (refs in [7]), a casein kinase 1 that phosphorylates tobacco mosaic virus (TMV) MP [8], a β-1,3-glucanase [9], and class 1 reversibly glycosylated proteins (C1RGPs) [10]. The lack of identification of the constituent components of plasmodesmata has remained an outstanding challenge in plant biology and has hindered a fuller understanding of the non–cell-autonomous control of plant development and the processes of tissue invasion by pathogens.

Bottom Line: We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking.These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication.They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.

View Article: PubMed Central - PubMed

Affiliation: John Innes Centre, Norwich Research Park, Colney, Norwich, United Kingdom.

ABSTRACT
Plasmodesmata provide the cytoplasmic conduits for cell-to-cell communication throughout plant tissues and participate in a diverse set of non-cell-autonomous functions. Despite their central role in growth and development and defence, resolving their modus operandi remains a major challenge in plant biology. Features of protein sequences and/or structure that determine protein targeting to plasmodesmata were previously unknown. We identify here a novel family of plasmodesmata-located proteins (called PDLP1) whose members have the features of type I membrane receptor-like proteins. We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking. PDLP1a is targeted to plasmodesmata via the secretory pathway in a Brefeldin A-sensitive and COPII-dependent manner, and resides at plasmodesmata with its C-terminus in the cytoplasmic domain and its N-terminus in the apoplast. Using a deletion analysis, we show that the single transmembrane domain (TMD) of PDLP1a contains all the information necessary for intracellular targeting of this type I membrane protein to plasmodesmata, such that the TMD can be used to target heterologous proteins to this location. These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication. They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.

Show MeSH