As an obligate intracellular protozoan,Toxoplasma gondii is a successful pathogen infecting a variety of animals and humans.During host infection,micronemal proteins(MICs)are released to parasite surface,where they bind host cells to mediate gliding motility and host-cell attachment.As an adhesin,the micronemal protein MIC3 plays important roles in host cell attachment,as well as modulation of host EGFR signaling cascade.However,the specific host proteins that interact with MIC3 are unknown and the identification of such proteins will increase our understanding of how MIC3 exerts its functions.In this study,we sought to identify host proteins interacting with MIC3 by yeast two-hybrid screens.Using MIC3 as bait,we screened a library expressing mouse proteins and found five mouse proteins that showed positive interactions with MIC3.Two of which,spermatogenesis-associated protein 3(Spata3)and dickkopf-related protein 2(Dkk2),were further confirmed to interact with MIC3 both in vitro and in vivo by additional protein-protein interaction tests.We also found that the tandem repeat EGF domains of MIC3 were critical in mediating the interactions with the identified host proteins.This is the first study to show that MIC3 interacts with host proteins that are involved in reproduction,growth and development.The results will help us to better understand the functions of adhesion-associated micronemal proteins in Toxoplasma gondii.