Dukes was born in San Francisco, California. He had a long career in films, appearing in 35; and as a television guest star, notably as the man who attempted to rape Edith Bunker on All in the Family, and during the 1980s in the miniseries The Winds of War. He received an Emmy nomination for best supporting actor for his role in The Josephine Baker Story (1991) and appeared as Arthur Miller in Norma Jean & Marilyn (1996). He was also a regular in the first season of Sisters as the transvestite MD husband of oldest sister Alex (Swoosie Kurtz), and appeared occasionally as the character in subsequent seasons. On Dawson's Creek, he had a recurring role as Mr. McPhee, the father to Jack (Kerr Smith) and Andie (Meredith Monroe) from the second through fourth seasons.

Dukes had extensive stage experience, first appearing on Broadway in 1971 and later in a revival of Molière's The School for Wives. He also played such colorful roles as Dracula, Doctor Frankenstein, and Antonio Salieri in the original production of Amadeus - replacing Ian McKellan. He won plaudits when he replaced John Lithgow in the original production of David Henry Hwang's play M. Butterfly and a Tony nomination in 1980 for best featured actor in a play for Bent.

Dukes died in Spanaway, Washington while on location shooting the Stephen King mini-series Rose Red. His widow's experiences with the Pierce County, Washington medical examiner's office were so frustrating that she publicly spoke out against the office's lack of cooperation. The ME's office would not let her see her husband's body, did not inform her of her rights or honor some of the rights she did try to exercise, and failed to detect during his autopsy the evidence of a previous heart attack.

David Dukes was interred in the Forest Lawn Memorial Park Cemetery in Glendale, California.