Miles Morales, played by Shameik Moore, is a young teenager with a deep admiration for Spider-Man, yet he finds himself struggling to meet his parents' expectations. His father, Jefferson Davis, a dedicated police officer, views Spider-Man as a threat, which adds to Miles' inner turmoil. Living with Jefferson and his mother, Rio (Lauren Vélez), Miles navigates the challenges of adolescence while recently transferring to a prestigious school where he feels out of place. The only friend he makes there is Gwanda (Hailee Steinfeld), another new student who becomes his ally.
In secret, Miles ventures to visit his Uncle Aaron (Mahershala Ali), known as The Prowler, in an abandoned subway station, where he is introduced to the world of graffiti art. It is here that Miles is unexpectedly bitten by a radioactive spider, resulting in astonishing spider-like powers: he can turn invisible and unleash electric venom blasts. As he delves into the comic books, he learns that he may not be alone in this extraordinary journey as he discovers the existence of another Spider-Man.
Determined to understand more, Miles returns to the subway station, where he accidentally stumbles upon a particle accelerator engineered by the ominous Wilson Fisk, aka The Kingpin. Fisk's aim is to breach parallel universes in hopes of retrieving lost versions of his wife and son. During a fierce battle with the enforcers, including the Green Goblin and his Uncle Aaron, Miles witnesses a catastrophic event: the death of Spider-Man (Chris Pine), who sacrifices himself to save Miles and shares crucial information about the USB device designed to disable the accelerator.
With Spider-Man's legacy behind him, news of his death ripples across the city, and the particle accelerator inadvertently causes characters and elements from alternate universes to invade Miles's world. Despite donning a Spider-Man costume, Miles struggles to harness his newfound abilities and accidentally damages the vital USB drive. At Spider-Man's grave, he crosses paths with Peter B. Parker (Jake Johnson), an older, weary version of Spider-Man from another dimension, who reluctantly becomes his mentor after realizing he needs Miles' help to return to his own reality.
Peter B. Parker engages Miles in training, imparting wisdom on how to navigate his powers, but he is conflicted about Miles' inexperience. During a mission to infiltrate Fisk's facility, they encounter the cunning Olivia Octavius, who reveals the dire consequences of activating the particle accelerator again. Octavius, however, aligns herself with Kingpin's malevolent agenda.
Alongside them is Gwen Stacy, another version of a web-slinger, who joins the duo in a collective effort to enlist the help of other dimension-traveling heroes, including Spider-Man Noir, Spider-Ham, and Peni Parker, all of whom face threats due to their origin in different realities.
As they work toward disabling the accelerator, Miles' inexperience leads to doubt within the group, resulting in him retreating and uncovering his uncle's true identity as The Prowler. Tragically, this leads to a series of confrontations that culminate with Miles revealing himself to Aaron, which prompts a tragic showdown with Fisk.
The stakes rise as Miles rallies the diverse team of heroes, and they engage in a perilous battle against Kingpin and his ruthless associates. With newfound determination sparked by his father's recognition of Spider-Man's true nature, Miles finally embraces his destiny. Utilizing all he has learned, he battles Fisk and ultimately destroys the accelerator.
In the aftermath, the authorities take Fisk into custody, and the recognition of Spider-Man as a hero begins to blossom. Miles accepts the weight of responsibility that comes with his powers. Meanwhile, back in their respective dimensions, Peter B. Parker seeks to mend his relationship with Mary Jane, while Gwen devises a way to keep in touch with Miles across the multiverse. In another timeline, Miguel O'Hara finds himself drawn into a new crisis, armed with advanced dimension-hopping technology, setting the stage for further adventures.
