Molecular Mechanism Of TgATG12 Promoting The Localization Of TgATG8 On The Membrane Of Apicoplast In Toxoplasma Gondii

Toxoplasmosis is a zoonotic parasitic disease with high infection rate,and the causing pathogen is Toxoplasma gondii.T.gondii belongs to Apicomplexa,and contains a plastid homologous to chloroplasts of plant and algae.Apicoplast plays an important role in the survival of T.gondii.Recent studies have shown that TgATG8 is partially located to the outer membrane of the apicoplast in T.gondii.Deletion of TgATG8 and other autophagy related genes,such as TgATG4 and TgPROP2,which regulate TgATG8 conjugation in macroautophagy,lead to the loss of the apicoplast and in turn affect the survival of T.gondii.In classical autophagy,ATG5-ATG12-ATG16 complex acts like an E3 ubiquitin ligase and plays a crucial role in the conjugation of ATG8 onto PE.However,the molecular mechanism of TgATG8 conjugation on the apicoplast of T.gondii is still unclear.In this study,we inserted a SmFP-Myc tag at the N-terminal of TgATG12 to analyze the subcellular location of TgATG12.Then,a conditional knockout strain was constructed based on a auxin-inducible degron(AID)system to analyze the role of TgATG12 on the apicoplast biogenesis of T.gondii.The results showed that TgATG12 localized in the cytoplasm of T.gondii and accumulated at a adjacent area of apicoplast.Depletion of TgATG12 resulted in the inability of TgATG8 conjunct to PE,and the loss of the apicoplast and ultimately affected the lytic cycle of T.gondii.In summary,the molecular mechanism of TgATG12 promoting the association of TgATG8 with the outer membrane of T.gondii apicoplast was preliminarily analyzed using AID-tagging method.Our research laid a foundation for the further investigation of the molecular mechanism of location of TgATG8 at the outer membrane of the apicoplast.