After attending private boarding school at Culver Military Academy in Culver, Indiana, Winters enlisted in the Marines at age 17 and served in the South Pacific during World War II. After his discharge, he studied cartooning at Dayton Art Institute, where he met Eileen Schauder, whom he married in 1948.
As a stand-up comic with a madcap wildness, Winters recorded many classic comedy albums for the Verve Records label, starting in 1960. Probably the best-known of his characters from this period is Maude Frickert, the seemingly sweet old lady with the barbed tongue. He was a favorite of Jack Paar and appeared frequently on his television programs. In addition, he would often appear on the Tonight Show with Johnny Carson, usually in the guise of some character. Carson often did not know what Winters had planned and usually had to tease out the character's back story through the course of the interview.
Winters has appeared in nearly 50 movies and several television shows, including particularly notable roles in the film It's a Mad, Mad, Mad, Mad World and in the dual roles of Henry Glenworthy and his dark, scheming brother, the Rev. Wilbur Glenworthy, in the film adaptation of Evelyn Waugh's The Loved One. Fellow comedians who starred with him in "Mad World," such as Arnold Stang, claimed that in the long periods while they waited between scenes, Winters would entertain them for hours in their trailer by becoming any character that they would suggest to him.
On television, in the late sixties, he appeared as a regular (along with Woody Allen and Jo Anne Worley) on the Saturday morning children's program Hot Dog. In the seventies, he appeared in his own show, The Wacky World of Jonathan Winters (1972–74). Winters has also done some dramatic work, as evidenced in The Twilight Zone episode "A Game of Pool" (episode #3.5, 13 October, 1961). He recorded Ogden Nash's The Carnival of the Animals poems to Camille Saint-Saëns' classical opus. He also made an appearance on a Dean Martin Comedy Roast. Winters appeared on ABC's The American Sportsman, hosted by Grits Gresham, who took celebrities on hunting, fishing, or shooting trips to exotic places around the world.
In the fourth and last season of the sci-fi-based TV comedy Mork & Mindy, Jonathan Winters (one of Robin Williams' idols) was brought in as Mork & Mindy's child, Mearth. Due to the different Orkan physiology, Mork laid an egg, which grew and hatched into the much older Winters. It had been previously explained that Orkans aged "backwards," thus explaining Mearth's appearance and that of his teacher, Miss Geespot (portrayed by then-11-year-old actress Luanne). Mork's infant son Mearth in Mork & Mindy was created in hopes of improving ratings and as an attempt to capitalize on Williams' comedic talents. Winters had previously guest starred in Season 3, Episode 18 as Dave McConnell, Mindy's Uncle. Although Robin Williams calls Jonathan Winters his greatest influence, the idea of Mearth didn't work, and the show was soon canceled, in 1982.
In 1991-1992 he was on Davis Rules, a sitcom that lasted two seasons(25 episodes). He played Gunny Davis, an eccentric grandfather that was helping raise his grandchildren after his son had lost his wife. In addition to his live action roles, he was also a guest star on The New Scooby-Doo Movies as a sweet old lady who was really the villain, and was the narrator in Frosty Returns. Winters had also earlier appeared as himself on an episode of Scooby-Doo, where the Scooby Gang was looking forward to his promised performance as Maude Frickert. Along with numerous roles in Scooby-Doo, Winters also provided the voice for the thief in The Thief and the Cobbler (Miramax version).
From 1959 to 1964, Winters' voice could be heard in a series of popular television commercials for Utica Club beer. In the ads, he provided the voices of talking beer steins, named "Shultz and Dooley." Later, he became a spokesman for Hefty brand trash bags—for whom he appeared as a dapper garbageman known for collecting "gahr-bahj," as well as Maude Frickert and other characters.
In June 2008, Winters was presented with the TV Land Pioneer Award by his friend Robin Williams.