More than 30 years after graduating from Top Gun,[a] United States Navy Captain Pete "Maverick" Mitchell is a test pilot. Despite many achievements, repeated insubordination has kept him from flag rank;[b] his friend and former Top Gun rival, Admiral Tom "Iceman" Kazansky, now Commander of the U.S. Pacific Fleet, often protects Maverick. Rear Admiral Chester "Hammer" Cain plans to cancel Maverick's hypersonic "Darkstar" scramjet program in favor of funding drones. To save the program, Maverick unilaterally changes the target speed for that day's test from Mach 9 to the final contract specification of Mach 10. However, the prototype is destroyed when he cannot resist pushing beyond Mach 10. Iceman again saves Maverick's career by assigning him to the Top Gun school at NAS North Island for his next assignment, but Cain tells Maverick that the era of crewed fighter aircraft will soon be over.

The Navy has been tasked with destroying an unsanctioned uranium enrichment plant, located in an underground bunker at the end of a canyon, before it becomes operational. It is defended by surface-to-air missiles (SAMs), GPS jammers, and fifth-generation Su-57 fighters as well as older F-14 Tomcats. Maverick devises a plan employing two pairs of F/A-18E/F Super Hornets armed with laser-guided bombs, but instead of participating in the strike, he is to train an elite group of Top Gun graduates assembled by Air Boss Vice Admiral Beau "Cyclone" Simpson.

Maverick dogfights his skeptical students and prevails in every contest, winning their respect. Lieutenants Jake "Hangman" Seresin and Bradley "Rooster" Bradshaw—son of Maverick's deceased best friend and RIO Nick "Goose" Bradshaw—clash: Rooster dislikes Hangman's cavalier attitude, while Hangman criticizes Rooster's cautious flying. Maverick reunites with former girlfriend Penny Benjamin, to whom he reveals that he promised Rooster's dying mother that Rooster would not become a pilot. Rooster, unaware of the promise, angrily resents Maverick for dropping his Naval Academy application—impeding his military career—and blames him for his father's death. Maverick is reluctant to further interfere with Rooster's career, but the alternative is to send him on the extremely dangerous mission. He tells his doubts to Iceman, who has terminal throat cancer. Iceman advises that "It's time to let go" and reassures him that both the Navy and Rooster need Maverick.

Iceman dies not long after, and Cyclone removes Maverick as instructor following a training incident in which an F/A-18F is lost. Cyclone relaxes the mission parameters, so they are easier to execute but make escape much more difficult. During Cyclone's announcement, Maverick makes an unauthorized flight through the course with his preferred parameters, proving that it can be done. Cyclone reluctantly appoints Maverick as team leader.

Maverick flies the lead F/A-18E, accompanied by a buddy lazing F/A-18F[c] flown by Lieutenant Natasha "Phoenix" Trace and WSO Lieutenant Robert "Bob" Floyd. Rooster leads the second strike pair, which includes Lieutenant Reuben "Payback" Fitch and WSO Lieutenant Mickey "Fanboy" Garcia. The four jets launch from an aircraft carrier, and Tomahawk cruise missiles destroy the nearby air base as they approach. The teams destroy the plant, but the SAMs open fire during their escape, as anticipated. Rooster runs out of countermeasures, and Maverick sacrifices his plane to protect him. Believing Maverick to be dead, the others are ordered back to the carrier, but Rooster returns to find that Maverick ejected and is being targeted by an Mi-24 attack helicopter. After destroying the gunship, Rooster is shot down by a SAM and ejects. The two rendezvous and steal an F-14 from the damaged air base. Maverick and Rooster destroy two intercepting Su-57s, but a third attacks as they run out of ammunition and countermeasures. Hangman unexpectedly arrives in time to shoot it down, and the planes return safely.

Later, Rooster helps Maverick work on his P-51 Mustang. Rooster looks at a photo of their mission's success, pinned alongside a photo of his late father and a young Maverick, as Penny and Maverick fly off in the P-51.