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The plasmodesmal protein PDLP1 localises to haustoria-associated membranes during downy mildew infection and regulates callose deposition.

Caillaud MC, Wirthmueller L, Sklenar J, Findlay K, Piquerez SJ, Jones AM, Robatzek S, Jones JD, Faulkner C - PLoS Pathog. (2014)

Bottom Line: We found that the pdlp1,2,3 triple mutant is more susceptible to Hpa while overexpression of PDLP1 enhances plant resistance, suggesting that PDLPs enhance basal immunity against Hpa.Haustorial encasements are depleted in callose in pdlp1,2,3 mutant plants whereas PDLP1 over-expression elevates callose deposition around haustoria and across the cell surface.These data indicate that PDLPs contribute to callose encasement of Hpa haustoria and suggests that the deposition of callose at haustoria may involve similar mechanisms to callose deposition at plasmodesmata.

View Article: PubMed Central - PubMed

Affiliation: The Sainsbury Laboratory, Norwich Research Park, Norwich, United Kingdom.

ABSTRACT
The downy mildew pathogen Hyaloperonospora arabidopsidis (Hpa) is a filamentous oomycete that invades plant cells via sophisticated but poorly understood structures called haustoria. Haustoria are separated from the host cell cytoplasm and surrounded by an extrahaustorial membrane (EHM) of unknown origin. In some interactions, including Hpa-Arabidopsis, haustoria are progressively encased by host-derived, callose-rich materials but the molecular mechanisms by which callose accumulates around haustoria remain unclear. Here, we report that PLASMODESMATA-LOCATED PROTEIN 1 (PDLP1) is expressed at high levels in Hpa infected cells. Unlike other plasma membrane proteins, which are often excluded from the EHM, PDLP1 is located at the EHM in Hpa-infected cells prior to encasement. The transmembrane domain and cytoplasmic tail of PDLP1 are sufficient to convey this localization. PDLP1 also associates with the developing encasement but this association is lost when encasements are fully mature. We found that the pdlp1,2,3 triple mutant is more susceptible to Hpa while overexpression of PDLP1 enhances plant resistance, suggesting that PDLPs enhance basal immunity against Hpa. Haustorial encasements are depleted in callose in pdlp1,2,3 mutant plants whereas PDLP1 over-expression elevates callose deposition around haustoria and across the cell surface. These data indicate that PDLPs contribute to callose encasement of Hpa haustoria and suggests that the deposition of callose at haustoria may involve similar mechanisms to callose deposition at plasmodesmata.

No MeSH data available.


Related in: MedlinePlus

PDLPs localise to the extra-haustorial membrane.PDLP1-GFP is observed at the EHM prior to encasement. (A) In unencased haustoria, or those with a developing encasement at the haustorial neck, PDLP1-GFP is present in the EHM surround the haustorium (n = 50/50). PD-located signal is indicated with solid arrows, while open arrows indicate signal associated with haustoria. (B) As the encasement develops (stained with aniline blue, open arrows) and surrounds the entire haustorium PDLP1-GFP fluorescence remains associated with the haustorium and PDLP1-GFP positive bodies can be seen at the encasement periphery (see E). (C) When encasements are mature PDLP1-GFP is no longer associated with the structure. (D) Scheme of the development of the Hpa haustorial encasement, stages I, II and III are indicated in (A–C) for reference. (E) Enlargement of developing encasements shows PDLP1-GFP positive bodies at the periphery of the encasement (arrows). Scale bars are 20 µm (A–C) and 10 µm (E).
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ppat-1004496-g002: PDLPs localise to the extra-haustorial membrane.PDLP1-GFP is observed at the EHM prior to encasement. (A) In unencased haustoria, or those with a developing encasement at the haustorial neck, PDLP1-GFP is present in the EHM surround the haustorium (n = 50/50). PD-located signal is indicated with solid arrows, while open arrows indicate signal associated with haustoria. (B) As the encasement develops (stained with aniline blue, open arrows) and surrounds the entire haustorium PDLP1-GFP fluorescence remains associated with the haustorium and PDLP1-GFP positive bodies can be seen at the encasement periphery (see E). (C) When encasements are mature PDLP1-GFP is no longer associated with the structure. (D) Scheme of the development of the Hpa haustorial encasement, stages I, II and III are indicated in (A–C) for reference. (E) Enlargement of developing encasements shows PDLP1-GFP positive bodies at the periphery of the encasement (arrows). Scale bars are 20 µm (A–C) and 10 µm (E).

Mentions: Given that PDLP1 is specifically expressed in haustoria-containing cells we examined the subcellular location of PDLP1-GFP after Hpa infection to determine if this increase in expression is likely to affect PD function. Plants that constitutively express PDLP1-GFP under the 35S promoter (PDLP1 OE, [33]) were imaged at 3–6 DPI to observe haustoria at various stages of encasement (Figure 2). In uninfected leaves, PDLP1-GFP localises to PD (white arrows, Figure 3A, [33]). Following inoculation with Hpa PDLP1-GFP was visible in the PD and surrounding unencased haustoria (Figure 2A, 3A). Infiltration of infected tissue with aniline blue stained any developing, callose-filled encasements. Haustoria with developing encasements showed aniline blue staining at the neck of the haustorium while PDLP1-GFP completely surrounded the structure (Figure 2A), illustrating that PDLP1-GFP associates with haustoria prior to development of the encasement. PDLP1-GFP remained associated with the haustorium as the encasement developed (Figure 2B) but was not associated with fully encased haustoria at a late stage of development (Figure 2C). During the encasement process, small PDLP1-GFP containing bodies could be seen peripheral to the haustorium (Figure 2E). These bodies are possibly secretory vesicles depositing encasement material at the developing structure. The localisation of the PDLP1-GFP fusion during infection was also imaged when expressed from its native promoter. In plants stably expressing PDLP1pro::PDLP1-GFP[33], PDLP1-GFP was observed surrounding haustoria (Figure S2A). Like in PDLP1 OE plants, PDLP1-GFP was also observed in the developing encasement, but sometimes this association with the encasement could be resolved into two layers that suggest PDLP1-GFP is concentrated in membranes surrounding the encasement (Figure S2B). Many PM proteins are not present in the EHM but are associated with the haustorial encasement, i.e. they are observed at the neck of haustoria early in encasement development and completely surrounding the haustorium when the encasement is fully developed [3]. Localisation of PDLP1 at haustorial membranes prior to encasement suggests it is differentially incorporated into the EHM relative to other PM proteins. Significantly, this localisation also indicates that PDLP1 has a non-PD associated function.


The plasmodesmal protein PDLP1 localises to haustoria-associated membranes during downy mildew infection and regulates callose deposition.

Caillaud MC, Wirthmueller L, Sklenar J, Findlay K, Piquerez SJ, Jones AM, Robatzek S, Jones JD, Faulkner C - PLoS Pathog. (2014)

PDLPs localise to the extra-haustorial membrane.PDLP1-GFP is observed at the EHM prior to encasement. (A) In unencased haustoria, or those with a developing encasement at the haustorial neck, PDLP1-GFP is present in the EHM surround the haustorium (n = 50/50). PD-located signal is indicated with solid arrows, while open arrows indicate signal associated with haustoria. (B) As the encasement develops (stained with aniline blue, open arrows) and surrounds the entire haustorium PDLP1-GFP fluorescence remains associated with the haustorium and PDLP1-GFP positive bodies can be seen at the encasement periphery (see E). (C) When encasements are mature PDLP1-GFP is no longer associated with the structure. (D) Scheme of the development of the Hpa haustorial encasement, stages I, II and III are indicated in (A–C) for reference. (E) Enlargement of developing encasements shows PDLP1-GFP positive bodies at the periphery of the encasement (arrows). Scale bars are 20 µm (A–C) and 10 µm (E).
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ppat-1004496-g002: PDLPs localise to the extra-haustorial membrane.PDLP1-GFP is observed at the EHM prior to encasement. (A) In unencased haustoria, or those with a developing encasement at the haustorial neck, PDLP1-GFP is present in the EHM surround the haustorium (n = 50/50). PD-located signal is indicated with solid arrows, while open arrows indicate signal associated with haustoria. (B) As the encasement develops (stained with aniline blue, open arrows) and surrounds the entire haustorium PDLP1-GFP fluorescence remains associated with the haustorium and PDLP1-GFP positive bodies can be seen at the encasement periphery (see E). (C) When encasements are mature PDLP1-GFP is no longer associated with the structure. (D) Scheme of the development of the Hpa haustorial encasement, stages I, II and III are indicated in (A–C) for reference. (E) Enlargement of developing encasements shows PDLP1-GFP positive bodies at the periphery of the encasement (arrows). Scale bars are 20 µm (A–C) and 10 µm (E).
Mentions: Given that PDLP1 is specifically expressed in haustoria-containing cells we examined the subcellular location of PDLP1-GFP after Hpa infection to determine if this increase in expression is likely to affect PD function. Plants that constitutively express PDLP1-GFP under the 35S promoter (PDLP1 OE, [33]) were imaged at 3–6 DPI to observe haustoria at various stages of encasement (Figure 2). In uninfected leaves, PDLP1-GFP localises to PD (white arrows, Figure 3A, [33]). Following inoculation with Hpa PDLP1-GFP was visible in the PD and surrounding unencased haustoria (Figure 2A, 3A). Infiltration of infected tissue with aniline blue stained any developing, callose-filled encasements. Haustoria with developing encasements showed aniline blue staining at the neck of the haustorium while PDLP1-GFP completely surrounded the structure (Figure 2A), illustrating that PDLP1-GFP associates with haustoria prior to development of the encasement. PDLP1-GFP remained associated with the haustorium as the encasement developed (Figure 2B) but was not associated with fully encased haustoria at a late stage of development (Figure 2C). During the encasement process, small PDLP1-GFP containing bodies could be seen peripheral to the haustorium (Figure 2E). These bodies are possibly secretory vesicles depositing encasement material at the developing structure. The localisation of the PDLP1-GFP fusion during infection was also imaged when expressed from its native promoter. In plants stably expressing PDLP1pro::PDLP1-GFP[33], PDLP1-GFP was observed surrounding haustoria (Figure S2A). Like in PDLP1 OE plants, PDLP1-GFP was also observed in the developing encasement, but sometimes this association with the encasement could be resolved into two layers that suggest PDLP1-GFP is concentrated in membranes surrounding the encasement (Figure S2B). Many PM proteins are not present in the EHM but are associated with the haustorial encasement, i.e. they are observed at the neck of haustoria early in encasement development and completely surrounding the haustorium when the encasement is fully developed [3]. Localisation of PDLP1 at haustorial membranes prior to encasement suggests it is differentially incorporated into the EHM relative to other PM proteins. Significantly, this localisation also indicates that PDLP1 has a non-PD associated function.

Bottom Line: We found that the pdlp1,2,3 triple mutant is more susceptible to Hpa while overexpression of PDLP1 enhances plant resistance, suggesting that PDLPs enhance basal immunity against Hpa.Haustorial encasements are depleted in callose in pdlp1,2,3 mutant plants whereas PDLP1 over-expression elevates callose deposition around haustoria and across the cell surface.These data indicate that PDLPs contribute to callose encasement of Hpa haustoria and suggests that the deposition of callose at haustoria may involve similar mechanisms to callose deposition at plasmodesmata.

View Article: PubMed Central - PubMed

Affiliation: The Sainsbury Laboratory, Norwich Research Park, Norwich, United Kingdom.

ABSTRACT
The downy mildew pathogen Hyaloperonospora arabidopsidis (Hpa) is a filamentous oomycete that invades plant cells via sophisticated but poorly understood structures called haustoria. Haustoria are separated from the host cell cytoplasm and surrounded by an extrahaustorial membrane (EHM) of unknown origin. In some interactions, including Hpa-Arabidopsis, haustoria are progressively encased by host-derived, callose-rich materials but the molecular mechanisms by which callose accumulates around haustoria remain unclear. Here, we report that PLASMODESMATA-LOCATED PROTEIN 1 (PDLP1) is expressed at high levels in Hpa infected cells. Unlike other plasma membrane proteins, which are often excluded from the EHM, PDLP1 is located at the EHM in Hpa-infected cells prior to encasement. The transmembrane domain and cytoplasmic tail of PDLP1 are sufficient to convey this localization. PDLP1 also associates with the developing encasement but this association is lost when encasements are fully mature. We found that the pdlp1,2,3 triple mutant is more susceptible to Hpa while overexpression of PDLP1 enhances plant resistance, suggesting that PDLPs enhance basal immunity against Hpa. Haustorial encasements are depleted in callose in pdlp1,2,3 mutant plants whereas PDLP1 over-expression elevates callose deposition around haustoria and across the cell surface. These data indicate that PDLPs contribute to callose encasement of Hpa haustoria and suggests that the deposition of callose at haustoria may involve similar mechanisms to callose deposition at plasmodesmata.

No MeSH data available.


Related in: MedlinePlus