Over 10,000 years before the events of Dune, sisters Valya and Tula Harkonnen join the Sisterhood, an order of women trained to serve the Great Houses of the Imperium as Truthsayers. Mother Superior Raquella Berto-Anirul dies after envisioning a future reckoning (Tiran-Arafel), which she dubs the Burning Truth. Valya uses the Voice to force Dorotea, Raquella's granddaughter/successor, to kill herself before she can destroy the Sisterhood's breeding program. Thirty years later, Valya, now Mother Superior, arranges for Princess Ynez, daughter of Emperor Javicco Corrino, to marry nine-year-old Pruwet Richese, which they expect will stabilize the Corrino bloodline for generations to come while strengthening their control of desert planet Arrakis and its invaluable spice. Imperial Truthsayer Kasha Jinjo has unsettling visions after meeting soldier Desmond Hart, who claims Imperium-based insurgents, and not the native Fremen, were responsible for the ambush that wiped out his regiment. Valya dismisses Kasha's concerns. Visiting a nightclub, Ynez has sex with Imperial Swordmaster Keiran Atreides. Following a conversation with Javicco that reveals his unease with Ynez's marriage, Desmond meets with Pruwet and uses a "great power" to immolate him, an attack that simultaneously immolates Kasha. Seeing Kasha's burning body prompts Valya to recall Raquella's vision.

After Kasha's inconclusive autopsy, Valya leaves with Theodosia to secure Princess Ynez. Natalya angers Ferdinand by claiming Pruwet's death is accidental. Desmond, who claims a sandworm swallowed him, tells Javicco he killed Pruwet for him; disturbed, Javicco has him arrested, but Natalya suggests they use Desmond. Valya has Tula put her protégée, Raquella's great-great-granddaughter Lila, through the Agony ritual to unlock her Other Memory and contact Raquella. Lila learns of a "revenant", envisions Valya killing her grandmother Dorotea, becomes overwhelmed, and seemingly dies. Constantine has sex with Shannon Richese and reveals Desmond's involvement in Pruwet's murder. Valya brings news of Kasha's death, agonizing Ynez, and interrogates Desmond, who again admits both murders; Javicco eschews prosecution. Keiran, a clandestine anti-imperial insurgent, copies the palace layout for the rebels and Ynez recounts how rebels once kidnapped her and Constantine. Ferdinand threatens Javicco and Desmond burns him as a warning. Valya acquires from her spy, Division bartender Sister Mikaela, a list of Arrakis rebels the Sisterhood secretly supports, saying she may surrender them to bolster Javicco. Desmond confronts Valya, saying he intends to destroy the Sisterhood. She employs the Voice to compel him to commit suicide but he successfully resists; Valya, shocked, retreats.

In the past, Valya advocates revenge on the Atreides for House Harkonnen's banishment. Her brother Griffin joins the Landsraad, seeking commercial privileges, but dies, allegedly murdered by Vorian Atreides. Valya joins the Sisterhood, and Tula, hiding her Harkonnen identity, visits lover Orry Atreides' Caladan bull-hunt. Dorotea criticizes Valya for questioning if Sisters can withhold truth from rulers, and for being without loyalties. At a rainy vow-taking ceremony, Valya refuses to put the Sisterhood first. She uses the Voice on Raquella, impressing her. Orry proposes to Tula; she accepts, and they have sex. Next morning, Tula has slaughtered his family, and kills him too, avenging Griffin. Raquella shows Valya her ambitious DNA database using forbidden technology. When Dorotea and Valya are to undergo the Agony, the latter refuses; one of Dorotea's followers calls her coward. Raquella remands Valya to Lankiveil, telling her to return as a Reverend Mother or not at all. During a tense exchange, Valya employs the Voice on her mother. She later puts herself through Agony, surviving; Tula joins her in the Sisterhood. In the present, Valya visits her family as a recourse; Tula pretends to euthanize a comatose Lila but connects her to a thinking/life-support machine.

Jen discovers other Sisters' nightmares, preventing Emeline's suicide. Valya becomes the Harkonnens' truthsayer. Ynez hears Desmond's murder confession. Desmond unsuccessfully recommends Javicco remove the Landsraad's Truthsayers. Keiran and Horace acquire an Ixian flying thermal bomb to kill Javicco, telling Mikaela. Valya hears, revealing she plans to save Javicco, and turn over Keiran. Sisters reveal Pruwet's murder to various Houses, which invite Harrow to replace the departed Richeses on the High Council and investigate. Tula has spice-imbibing acolytes draw images from their dreams: Shai-Hulud and blue eyes. Natalya reveals she leaked the truth about Pruwet's murder, so Javicco could prosecute thinking-machine abusers. At Division, Ynez recalls Vorian refused the throne, and tells Keiran of Desmond's guilt, volunteering to act. Harrow tells the Landsraad kanly was violated, almost naming the Corrinos; but Ynez accuses Desmond, who brings in Horace, bound. Desmond concurs but calls it a legal execution, flourishing the bomb. Javicco lets him immolate Horace and his co-conspirators. Valya obtains a sample of wounded Desmond's blood. Tula dreams she kills Emeline, who knows her crimes; Lila wakes. Evgeny laments Valya warping her siblings; criticizing his weakness, she withholds his respirator, killing him. Theodosia is revealed as a shapeshifter; Valya has her appear as Griffin.

Javicco makes Desmond an elite regiment's Bashar. Sister Francesca arrives to see Constantine, her son. Jen, trusted for her nightmare-resistance, is shown the ailing Lila. Harrow apologizes to Desmond for challenging Javicco, and blames Valya. Desmond wants proof of her complicity in rebel spice-smuggling. Harrow reports his meeting to Valya. Valya wants Francesca to assuage Javicco, and have Ynez study on Wallach IX. Raquella, possessing Lila, applauds Valya's efforts. Francesca tells Javicco to give Constantine responsibility; Javicco kisses her. Lila isolates a virus that killed Kasha; Avila, stunned, discovers Lila. Desmond's troops find spice at the Division; Keiran and Mikaela kill some. Keiran sees her Sisterhood knife, and warns her to avoid him. Desmond awakens after Mikaela's chain detonators explode the Division. Francesca says she had Constantine to protect Ynez. Valya gives Mikaela safe passage to Arrakis; Mikaela feels exploited. Constantine discovers Keiran's palace map, and smilingly betrays him to Desmond. Javicco makes Constantine fleet commander, tasked to pacify Arrakis. Ynez interrogates Keiran, who pledges to improve the Imperium. Tula is shocked to discover that Desmond has Atreides and Harkonnen blood; on Salusa Secundus, Desmond tells Natalya that his mother was a Sister, who left him to scavengers. Natalya complains to Desmond that the Sisterhood stole her power and her daughter, then kisses him.

In flashback, Tula sees Valya order Dorotea's suicide, then reveals her pregnancy. Valya shows Tula Anirul, and plans mating Javicco to Natalya. After birthing Desmond, Tula exchanges him for a stillborn. In the present, Nazir tells Tula fear feeds the virus, but dies analyzing it. Natalya arrests Ynez for attempting Keiran's rescue. Valya gives Francesca a poison needle to kill Javicco. Javicco unsuccessfully tries to send Natalya away. Javicco begs Valya's assistance against Desmond. She reveals the Sisterhood manipulated his whole life; he has her arrested. Dorotea-possessed Lila reveals Valya made Dorotea's followers commit suicide. Francesca shows angry Javicco the needle, blaming Valya. Valya frees Ynez and Keiran; they flee; Theodosia poses as Ynez, then a guard, stabbing Desmond. Javicco threatens Francesca, but, despairing, stabs himself. Natalya kills Francesca with the needle. Lila shows the acolytes Dorotea's followers' skeletons, then destroys Anirul. Harrow recovers a recording device he used to spy on Valya. Desmond ignites Valya's virus; Tula helps her to survive by abandoning fear. Valya senses an unknown person had a blue-"eyed" machine forcibly implant technology in Desmond. She considers killing Desmond, but Tula reveals his parentage. Valya says some hidden hand, grasping for control, also realized Desmond's potential. Tula embraces Desmond; his guards arrest her. Valya, Keiran, and Ynez land on Arrakis.