Stop! That! Train!

Best friends and train stewardesses Tess and DeeDee find themselves out of a job when their train line, the Stank Rail, is discontinued. They sneak onto the luxurious Glamazonian Express disguised as stewardesses, where they meet the arrogant Amber, Alli, and Ayshleiygh, who constantly upstage them.

Meanwhile, at the United States Train Command Center, employee Donna Dusk discovers that the Glamazonian Express is headed straight towards a massive storm, dubbed "Stormaganza". In the midst of the chaos, the Glamazonian Express loses their brakes; the train is headed for Florida and is expected to crash into a nuclear site situated between a shelter for abandoned dogs and the home of Laurie Metcalf. President Judy Gagwell receives the news and gives a statement, but suddenly begins to experience traumatic flashbacks of an incident where she accidentally ran over a little girl during her time in the discontinued military branch of United Train Force.

When the conductor of the Glamazonian Express is left incapacitated due to a scorpion bite, DeeDee and co-conductor Cal are left to get the train back on track, though they often find themselves distracted due to their growing attraction to one another. Due to the storm, they are forced to redirect the train into a haunted tunnel. Meanwhile, Tess finds herself assimilating well into the first-class culture, outshining and eventually replacing Amber. DeeDee is furious that Tess has abandoned her for the Glamazonian Express girls, while Tess angrily counters that DeeDee has been holding her back, causing a rift in the girls' friendship. After the tunnel incident, President Gagwell finds her approval ratings rapidly declining and steps down. Cal is accidentally sucked out of the train and falls down a ravine. Amber discovers the truth about Tess and DeeDee's past with Stank Rail and exposes them to the passengers. Donna is fired for the expected disaster and runs into Gagwell at a bar. After comiserating over their respective failures, a newly inspired Donna encourages Gagwell to take control of their fates and save the train.

President Gagwell boards the train while Dusk takes control of the command center. After Tess and DeeDee reconcile, they work together with Gagwell to eject the engine and stop the train before it runs into the Make-A-Wish foundation field trip to the Florida station. Afterwards, Cal is revealed to be alive; he and DeeDee profess their love to one another and kiss. Tess, DeeDee, and Donna are regarded as heroes by the media while President Gagwell returns to her position.