Uptake of oomycete RXLR effectors into host cells by clathrin-mediated endocytosis

ABSTRACT: Filamentous (oomycete and fungal) plant pathogens can deliver cytoplasmic effector proteins into host cells to facilitate disease. How RXLR effectors from the potato late blight pathogen Phytophthora infestans enter host cells is unknown. One possible route involves clathrin-mediated endocytosis (CME). Transient silencing of NbCHC, encoding clathrin heavy chain, or endosome marker NbAra6 in the model host Nicotiana benthamiana, attenuated P. infestans infection and reduced translocation of RXLR effector fusions from transgenic pathogen lines into host cells. In contrast, silencing PP1c isoforms, susceptibility factors that are not required for endocytosis, reduced infection but did not attenuate RXLR effector uptake. Endosome enrichment by ultracentrifugation and sucrose gradient fractionation revealed co-localisation of RXLR effector Pi04314-RFP with clathrin-coated vesicles. Immunopurification of clathrin- and NbAra6-associated vesicles during infection revealed that RXLR effectors Pi04314-RFP and AvrBlb1-RFP, but not apoplastic effector PiSCR74-RFP, were co-immunoprecipitated during infection with pathogen lines secreting these effectors. MS/MS analyses of proteins co-immunoprecipitated with NbAra6-GFP during infection revealed enrichment of host proteins associated with endocytic vesicles alongside multiple pathogen RXLR effectors, but not apoplastic effectors, including PiSCR74, which do not enter host cells. Our data show that uptake of P. infestans RXLR effectors into plant cells occurs via CME.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Phytophthora Infestans T30-4 Nicotiana Benthamiana

TISSUE(S): Plant Cell, Leaf, Haustorium

DISEASE(S): Disease Free

SUBMITTER: Haixia Wang  

LAB HEAD: Paul Birch

PROVIDER: PXD040285 | Pride | 2023-07-20

REPOSITORIES: Pride