Abstract
Eukaryotic cells deploy autophagy to eliminate invading microbes. In turn, pathogens have evolved effector proteins to counteract antimicrobial autophagy. How adapted pathogens co-opt autophagy for their own benefit is poorly understood. The Irish famine pathogen Phytophthora infestans secretes the effector protein PexRD54 that selectively activates an unknown plant autophagy pathway that antagonizes antimicrobial autophagy at the pathogen interface. Here, we show that PexRD54 induces autophagosome formation by bridging vesicles decorated by the small GTPase Rab8a with autophagic compartments labeled by the core autophagy protein ATG8CL. Rab8a is required for pathogen-triggered and starvation-induced but not antimicrobial autophagy, revealing specific trafficking pathways underpin selective autophagy. By subverting Rab8a-mediated vesicle trafficking, PexRD54 utilizes lipid droplets to facilitate biogenesis of autophagosomes diverted to pathogen feeding sites. Altogether, we show that PexRD54 mimics starvation-induced autophagy to subvert endomembrane trafficking at the host-pathogen interface, revealing how effectors bridge distinct host compartments to expedite colonization.
Highlights
Smaller autophagosomes of the cytosol-to-vacuole transport (Cvt) pathway do not rely on lipid droplets (LDs), suggesting that LDs are recruited for starvation-induced autophagy in order to meet the increased demand of lipids required for the biogenesis of larger sized autophagosomes (Dupont et al, 2014; Shpilka et al, 2015)
We have previously shown that ATG8 interacting motif (AIM) mediated binding of PexRD54 to ATG8CL is essential for the activation of autophagosome formation (Dagdas et al, 2016; Maqbool et al, 2016)
We reasoned that PexRD54 could stimulate autophagosome formation by either releasing the negative regulation of ATG8CL by host autophagy suppressors or through recruiting essential host components to the autophagosome biogenesis sites
Summary
Plant Material and Growth ConditionsN. benthamiana WT and transgenic plants (35S::GFP:Rab8a and 35S::GFP:ATG8CL) were grown and maintained in a greenhouse with high light intensity (16 hours light/8 hours dark photoperiod) at22-24°C. N. benthamiana WT and transgenic plants (35S::GFP:Rab8a and 35S::GFP:ATG8CL) were grown and maintained in a greenhouse with high light intensity (16 hours light/8 hours dark photoperiod) at. Plants were kept under a dark period of 24 hours before images were acquired. Images were acquired 3 days after infiltration (dpi). 35S::GFP:ATG8CL lines were produced as described elsewhere (1985) with the pK7WGF2::Rab8a and pK7WGF2::ATG8CL constructs, respectively. Cultures were grown and maintained by routine passing on rye sucrose agar medium at 18°C in the dark (van West et al, 1998). Zoospores were collected from 10-14 days old culture by flooding with cold water and incubation at 4°C for 90-120 minutes. Infection of agroinfiltrated leaves was carried out by addition of 10-mL droplets of zoospore solution at 50,000 spores/ml on detached N. benthamiana leaves (Chaparro-Garcia et al, 2011)
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