Her father (from whom she would become estranged due to his infidelities to her mother) insisted on the name Johnnie because he had wanted a boy, but she was often called Annie. She took up dancing to help exercise her legs to help her rickets.

She was considered a child dance prodigy. In an interview featured in a "behind the scenes" documentary on the making of the compilation That's Entertainment III, she claimed that Eleanor Powell was an early inspiration. Miller was given a contract with RKO at the tender age of thirteen (she had told them she was eighteen), and stayed there until 1940.

The following year, Miller was offered a contract at Columbia Pictures, where she bumped friend Lucille Ball from the throne as "Queen of the B-Movies". She finally hit her mark (starting in the late 1940s) in her roles in MGM musicals such as Kiss Me, Kate, Easter Parade, and On the Town.

Miller was famed for her speed in tap dancing; she claimed to be able to tap 500 times per minute. She was known as well, especially later in her career, for her distinctive appearance, which reflected a studio-era ideal of glamor: massive black bouffant hair, heavy makeup with a slash of crimson lipstick, and fashions that emphasized her lithe figure and long dancer's legs.

Her film career effectively ended in 1956 as the studio system lost steam to television, but she remained active in the theatre and on television. In 1979 she astounded audiences in the Broadway show Sugar Babies with fellow MGM veteran Mickey Rooney, which toured the United States extensively after its Broadway run. In 1983 she won the Sarah Siddons Award for her work in Chicago theatre.

She appeared in a special 1982 episode of The Love Boat, joined by fellow showbiz legends Ethel Merman, Carol Channing, Della Reese, Van Johnson, and Cab Calloway in a storyline that cast them as older relatives of the show's regular characters.

In 2001 she took her last role, playing Coco in auteur director David Lynch's critically acclaimed Mulholland Drive. Her last stage performance was a 1998 production of Stephen Sondheim's Follies, in which she played the hardboiled survivor Carlotta Campion and received rave reviews for her rendition of the anthemic "I'm Still Here."

For her contribution to the motion picture industry, Ann Miller has a star on the Hollywood Walk of Fame at 6914 Hollywood Blvd.

She died at the age of 80 from cancer that had spread to her lungs and was interred in the Holy Cross Cemetery in Culver City, California.