T. gondii is an obligate intracellular parasite, belonging to the Phylum Apicomplexa, infecting all warm-blooded animals including humans. During host cell invasion, specialized cytoskeletal and secretory organelles play a pivotal role. ROP18, as a member of the ROP2 family, has been identified as a key virulence factor mediating pathogenesis in T. gondii. Here, we identify an ER-resident protein, Derlin2, a factor implicated in the removal of misfolded proteins from the ER for cytosolic degradation, as a component of the machinery required for ER-associated protein degradation (ERAD). We identified Derlin2 interacting with ROP18 by yeast two-hybrid screening system. The interaction between ROP18 and Derlin2 was further confirmed through in vitro GST pull-down and in vivo immunoprecipitation assays. By immunofluorescence assay, we found that ROP18 co-localized with Derlin2 in the endoplasmic reticulum. Using overexpression and knockdown approaches, we demonstrated that Derlin2 was required for T. gondii ROP18 degradation. Consistently, cycloheximide chase experiments showed that the degradation of ROP18 relied on the Derlin2, but not Derlin1. These results indicate that interaction between Derlin2 and ROP18 is functionally relevant and leads ultimately to degradation of ROP18. The finding provides the basis for future studies on Derlin2-dependent ERAD of T. gondii ROP18 and subsequent antigen generation.
Animals Humans