DeLuise almost always appears in comedic parts, although an early appearance (in the movie Fail-Safe as a nervous enlisted soldier) showed a possible broader range. His first acting credit was as a regular performer in the television show The Entertainers in 1964. He has often co-starred with Burt Reynolds; together they starred in the film Cannonball Run. DeLuise was the host of the television show Candid Camera from 1991 to 1992. He is probably best known as a regular in Mel Brooks' films. Brooks' late wife, actress Anne Bancroft, directed Dom in Fatso (1980).

He has three sons, Peter, Michael, and David, all of whom are actors. Dom, his sons and his wife, actress Carol (to whom he has been married since 1965), occasionally appear in each others' projects.

He is sometimes credited as Dom De Luise, Dom Deluise, or Dom De Luises.