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Set one year after the events of the original series, Jeremie Belpois, Odd Della Robbia, Ulrich Stern, William Dunbar, Yumi Ishiyama and Aelita Schaeffer return to their daily lives and routines at Kadic Academy. X.A.N.A., despite their success in defeating it previously, suddenly reappears, reborn with even more strength and power than ever before. The protagonists resume their former double lives in order to protect humanity from X.A.N.A.’s evil schemes once again. Joined by William Dunbar, who has finally been accepted as the sixth Lyoko warrior, and an unreliable girl-genius named Laura Gauthier, the seven heroes are bent on unraveling the reasons for such a return and to exterminate X.A.N.A, the malevolently evil sentient, autonomous artificial intelligence/multi-agent system that is threatening the world once again.

In addition to X.A.N.A., the gang eventually discover that they have another dangerous enemy, a mad scientist called Professor Tyron who seems to be the one responsible for unintentionally returning X.A.N.A. to life. He commands a group of several virtual human avatars in green and black-striped costumes to counter the Lyoko warriors. In Tyron’s lab, the group also discovers Aelita’s long-lost mother, Anthea Hopper-Schaeffer. They seek to discover why she is with their new enemy and how to reunite Anthea and her daughter.

In the cliffhanger episode “Ultimate Mission”, it is revealed that Tyron had been married to Anthea for four years and is now Aelita’s stepfather and legal guardian. His only purpose of marrying Anthea was to get his hands on her late husband’s archives so that he could effectively create the Cortex region, which X.A.N.A. then secretly inhabited to carry out its plans for world domination and destruction once again.