The series serves as a conclusion to the storyline of Hope's Peak Academy, the elite school where the Super High School Level Despair, Junko Enoshima, played hostess to a "killing school life" among its students under the guise of the evil bear Monokuma.
Following the events of Danganronpa 2: Goodbye Despair, Makoto Naegi, the Super High School Level Hope, is placed under arrest by the Future Foundation for harboring the Remnants of Despair; those who continued the evil desires of the Super High School Level Despair, Junko Enoshima, following her death. The top members of the Future Foundation, including Makoto's former classmates Kyoko Kirigiri, Aoi Asahina, and Yasuhiro Hagakure, are all brought together to discuss Makoto's punishment. Just then, the Future Foundation's headquarters is attacked by a missile, cutting off all exits, before a round of sleeping gas knocks everyone unconscious. When they wake up, the members are greeted by the all-too-familiar visage of Monokuma, who announces that a new killing game has begun before revealing that housekeeper Chisa Yukizome has already been killed.
Monokuma reveals the rules of his new "Monokuma Hunter" game, in which each of the participants carry a wristband which, following a time limit, will put everyone to sleep for a short period of time. During this time, an unknown traitor will awaken and kill one of the other participants, meaning the participants must try to determine who the true attacker is and kill them before they kill everyone else. Additionally, if anyone attempts to remove the wristband or performs a personalised forbidden action, then they will be injected with a deadly poison. Therapist Miaya Gekkogahara attempts to hack Monokuma with her Usami program, but Monokuma hacks her right back and remodels Usami into Monomi. When boxer Juzo Sakakura starts acting violent against animator Ryota Mitarai, farmer Daisaku Bandai ends up triggering his forbidden action, witnessing violence by participants, and is killed by the poison in his wristband. As Juzo and student council president Kyosuke Munakata accuse Makoto of being the traitor and attempt to kill him, wrestler Great Gozu stands against them and escapes with Makoto, Aoi, and Miaya. However, after everyone falls asleep following the first time limit, Makoto wakes up to discover Aoi impaled with a knife.
Makoto is relieved to find Aoi is still alive, having only been depicted with a toy knife and some ketchup, only to discover Great Gozu had been killed instead. Hoping to prevent any further killings, Makoto uses the intercom to communicate with the other participants, explaining how he tried to help Remnants of Despair while also revealing his forbidden action. Meanwhile, pharmacist Seiko Kimura accuses sweetmaker Ruruka Ando of being the traitor, taking a bunch of drugs to transform herself and pursue her and blacksmith Sonosuke Izayoi. While Juzo goes after Kyoko, who is with Ryota and former headmaster Kazuo Tengan. Kyosuke, believing Makoto lacks the despair to make his words credible, attacks Makoto with his sword.
As Kyoko ends up spraining her ankle after Ryota lands on her, Kazuo manages to knock out Juzo by taking advantage of his inability to punch. Meanwhile, Kyosuke manages to capture Makoto, announcing his intention to execute him in order to lure out the attacker. He explains to Makoto that he suspects the traitor will end the killing game prematurely, frame one of the dead leaders for their crimes, and manipulate Makoto into destroying the Future Foundation. While Kazuo confronts Kyosuke, helping Aoi and Miaya escape with Makoto, Kyoko and Ryota, along with scoutman Koichi Kizakura, look over the corpses, pondering about the attacker's motives. Following a fierce battle, both Kyosuke and Kazuo end up falling off a walkway, and Kazuo is impaled on a piece of debris below. Meanwhile, Juzo makes his way past Miaya, unaware that she is actually a robot being controlled by the Lil Ultimate Homeroom, Monaca Towa.
As Seiko is confronted by Sonosuke and Ruruka, she recalls how her friendship with Ruruka became soured over the years. Just as Ruruka is cornered, she deduces Seiko's forbidden action, letting someone step on her shadow. Meanwhile, Juzo confronts Makoto and Aoi while Kazuo tells Kyosuke who attacker is, stabbing his eye before he finishes him off. When Yasuhiro, stuck outside of the building following the initial attack, is attacked by a helicopter, the explosion reveals a hidden secret entrance which Sonosuke discovers. Rejecting Ruruka's offer to kill Kyosuke in exchange for her life, Seiko takes more drugs and destroys all the lights in the room, pursuing Ruruka until she gets knocked out by Miaya's weapons. Regretting that she couldn't save anyone, Seiko is killed following the next time limit.
After looking over Great Gozu's body, Kyoko's group discover that, along with Seiko, Sonosuke has also been killed with a knife to the chest, soon coming across Ruruka. Meanwhile, Makoto notices that the rules don't forbid anyone from escaping, managing to get in contact with fellow classmate Byakuya Togami to tell them about Monokuma's killing game. Byakuya informs Makoto that an attack is being launched on Jabberwock Island, where the recovered Remnants of Despair are, while hiding the fact that the real Miaya Gekkogahara is dead. Believing that Monaca is involved, Byakuya gets in contact with Toko Fukawa and Makoto's sister Komaru in Towa City to gain their assistance.
Receiving help from the former Warriors of Hope, Komaru and Toko, along with the latter's psychotic alter-ego Genocider Sho, make their way to Monaca's hideout. After arriving at the scene and surviving an explosion, the girls discover Monaca has suddenly given up on despair, revealing that, outside of killing Miaya and replacing her with a robot, she is not the mastermind behind the new killing game. Leaving behind some disturbing words about Makoto, Monaca decides to launch herself into space to become a NEET, with Komaru preventing Toko from pursuing her any further. Afterwards, Komaru contacts Makoto and Aoi through the Miaya robot, relaying Monaca's warning that one of the original survivors of Hope's Peak Academy's mutual killing game will allegedly die because of Makoto.
As Byakuya arrives at the facility, managing to take out the helicopter that was pursuing Yasuhiro, the Miaya robot switches to automatic control and goes up against Kyosuke. Meanwhile, as Juzo prepares to attack Kyoko, Koichi, deducing that Juzo's forbidden action prevents him from punching anyone, lures him into a floor activated trap. Just then, Ruruka collapses the floor underneath Kyoko, prompting Koichi to break his forbidden action; opening his left hand, in order to save her, dying from poison as a result. After Ruruka fails to put Juzo under control with her sweets, Kyoko deduces that Ruruka is the one who killed Sonosuke, who actually died from his wristband's poison after eating one of her sweets. Kyoko then reveals the secret exit Sonosuke had found, deducing that Ruruka killed him to prevent him from triggering her own fobidden action; letting anyone escape from the game. Just as Byakuya manages to break into the facility, he and his team inadvertently set off a bomb that causes the facility to start collapsing.
The secret exit turns out to be a dead end, with Kyoko deducing the game area is actually somewhere underwater. After destroying the Miaya robot and retrieving a data module containing information on everyone's forbidden actions, Kyosuke suddenly turns against Juzo and kills him. Meanwhile, Makoto and Aoi meet up with Kyoko and Ryota before the next time limit, during which Ruruka is killed. Upon waking up, however, Makoto and the others are shocked to discover that Kyoko has died from her forbidden action, which was set to kill her if Makoto survived by the fourth time limit, at which point Kyosuke calls Makoto out.
Realising that Kyoko kept quiet about her forbidden action so that he could keep on living, Makoto leaves Aoi and Ryota with Kyoko as he sets off to confront Kyosuke. Kyosuke begins hunting Makoto throughout the facility, overcoming every trap that Makoto sets up for him. Using Kyosuke's forbidden action, opening doors, to keep him from killing him and becoming trapped, Makoto tries to talk with Kyosuke, who reveals Kazuo's final message that the attacker isn't merely one person, but in fact all of the Future Foundation members. Despite hearing this, Makoto states that even if it everyone were Remnants of Despairs, he would still feel glad to have met Kyoko, leading Kyosuke to realise just how much he cared for Chisa. As Kyosuke loses his will to keep fighting, Aoi appears with Kyoko's notebook, which allegedly reveals who the true attacker is.
Kyoko's notes reveal that everyone who was allegedly killed by the attacker actually committed suicide, with all their deaths taking place in rooms with monitors. In order to test this theory, Makoto has the others tie him up in a room with monitors before the next time limit. While the others sleep safely in another room, Makoto is awoken by a recording of Monokuma and shown a subliminal video that drives him crazy with visions of all of his dead classmates. Just as Makoto is brought to the brink of despair and prepares to kill himself, he is stopped by Juzo, who had cut off his own hand along with his bangle in order to observe the attacker. Learning what Makoto had disovered about the attacker, Juzo declines to return to the others with him, feeling guilty for betraying Kyosuke after being blackmailed by Junko. After Makoto reports his findings, Kyosuke tells the others that the game was likely set up by Kazuo, believing he was one of the Remnants of Despair. Using the last of his stength, Juzo brings himself to the power room to shut off power in the vicinity and disable the monitors and bangles, bleeding to death by the time Kyosuke finds him. Meanwhile, as Ryota is confronted by Makoto over the techniques used in Monokuma's video, he receives a message on his phone from Kazuo.
Kazuo's video message, sent to Ryota following his death, appears to prove he was a Remnant of Despair. Refusing to run away, Ryota explains how Junko stole his brainwashing techniques, intended for a peacemaking anime, to create a Despair Video, hypnotising people into becoming Remnants of Despair. He then reveals he has made a "Hope Video" possessing the opposite effect, using it on Aoi to have her hold Makoto back while he goes off on his own. Just as Makoto realises that Kazuo was actually trying to stop Ryota from using his video, the group are forced to defend themselves from soldiers hypnotised by Ryota's video. As Ryota escapes and prepares to broadcast his Hope Video worldwide, Makoto is aided by the arrival of Byakuya and Yasuhiro, while another figure makes his own move.