The program was an adventure series about Sergeant William Preston of the North-West Mounted Police and his lead sled dog, Yukon King, as they fought evildoers in the Northern wilderness during the Gold Rush of the 1890s. Preston, according to radio historian Jim Harmon, first joined the Mounties to capture his father's killer, and when he was successful he was promoted to sergeant. Preston worked under the command of Inspector Conrad, and in the early years was often assisted by a French-Canadian guide named Pierre.

Preston's staunchest ally, who was arguably the true star of the show and indeed often did more work than he did, was the brave Alaskan husky, Yukon King. Typical plots involved the pair helping injured trappers, tracking down smugglers, or saving cabin dwellers from wolverines. Sergeant Preston's faithful steed was Rex, used primarily in the summer months, but generally Yukon King and his dog team were the key mode of transportation (as signaled by Preston's cry of "On, King! On, you huskies!)."