In the novel, "Mr. Nancy"—an incarnation of the West African trickster god Anansi—dies, leaving twin sons, who in turn discover one another's existence after being separated as young children.

The reason Charlie had apparently not inherited any divine powers was because they had been passed down to his hitherto unknown brother, who Mrs. Higgler explains could be contacted by simply sending an invitation by talking to a spider.

Charlie is skeptical, and on his return to England largely forgets what Mrs. Higgler had told him, until one night when he drunkenly whispers to a spider that it would be nice if his brother stopped by for a visit.

The next morning the suave and well-dressed brother, going under the name of "Spider", visits Charlie and is shocked to learn that their father has died.

At Spider's recommendation, the two brothers attempt to drown their sorrows and become uproariously drunk on the proverbial trio of wine, women, and song.

While at work, Spider discovers his boss Grahame Coats's long-standing practice of embezzling from his clients and also steals the affection and virginity of Charlie's fiancée, Rosie Noah.

Embittered by the loss of his fiancée, Charlie uses his holiday to return to Florida and requests help from Mrs. Higgler and three of her equally old and eccentric friends to expel Spider.

Being themselves unable to banish Spider, they instead send Fat Charlie to "the beginning of the world," an abode of ancient gods similar to his father, each of whom represents a species of animal.

Finally, Charlie meets Bird Woman, who agrees to trade him her help, symbolised by one of her feathers, in exchange for "Anansi's bloodline".

Meanwhile, in London, a swindled client, Maeve Livingstone, confronts Grahame Coats directly, having learned of the theft of her late husband's royalties.

Maeve is contacted by her late husband, who advises her to move on to the afterlife, but she refuses in favour of taking vengeance on Grahame Coats.

The two discuss matters in the course of fleeing from birds from around the world, realising that giving away Anansi's bloodline implicates Charlie as well as Spider.

Meanwhile, Rosie and her mother have taken a consolation cruise to the Caribbean, where they unexpectedly meet Grahame Coats, who offers them a tour of his home.

Fat Charlie finally finds Mrs. Higgler after a long search in Saint Andrews and is sent again, via seance, to the beginning of the world where he forces the Bird Woman to give back Anansi's bloodline in return for her feather.

Tiger now takes possession of Grahame Coats's body and uses his bloodlust to manipulate him, intending to get revenge on Spider by killing Rosie and her mother.

Meanwhile, still at the beginning of the world, Charlie, having discovered his power to alter reality by singing a story, recounts the long tale of all that has gone before and humiliates Tiger to the point of retreating to his cave.

It stars Lenny Henry as Spider and Fat Charlie, Matt Lucas as Grahame Coats and Tiger, Rudolph Walker as Anansi, Doña Croll as Mrs Noah and the Bird Woman, Tameka Empson as Mrs Higgler, Petra Letang as Rosie, Jocelyn Jee Esien as Daisy, and Ben Crowe as Cabbies and other voices.

It is narrated by Joseph Marcell, and stars Jacob Anderson as Fat Charlie, Nathan Stewart-Jarrett as Spider and Lenny Henry as Anansi, with the rest of the cast including Julian Rhind-Tutt as Grahame Coats and Adjoa Andoh as Bird Woman.

In 2024, Gaiman partnered with Marc Bernardin to adapt Anansi Boys into a multi-volume comic series, with the artwork done by Shawn Martinbrough.