O'Brien, like Annette Funicello, was personally selected to audition for The Mickey Mouse Club by Walt Disney, in the spring of 1955. Disney had been alerted to him by a staff member, who caught his live performance at a charity gala.
Though he had little prior experience in singing or dancing, O'Brien was placed on The Mickey Mouse Club's first-string "Red Team" right from the start. He quickly picked up enough dance skills to perform in musical numbers, though his solo performances remained centered around his drums. He remained with the show for all three seasons (1955–1958) of original programming, and after filming stopped, went on live-performance tours with other Mouseketeers to Australia in 1959 and 1960.
Cubby was the only Mouseketeer allowed to play musical instruments on camera whether this had something to do with Walt casting him or not is unknown. Certainly Cubby never had the rumors of favoritism surrounding him like Annette did, possibly because his skill was obvious to all. In the show's first week, he got to play drums for Talent Round-Up day in a combo with his father Hack and brother Warren. From then on he was given many opportunities to play rhythm instruments, but his talent didn't end with music; he was a fine comedic actor, and learned enough dancing on the show to fake his way through the numbers. His singing was OK, though he wasn't called on for solos like Karen Pendleton.
Cubby appeared in the Disney film Westward Ho, the Wagons with Tommy Cole, Karen, and Doreen Tracey in 1956. The four Mouseketeers had barely any lines and very few scenes in this tepid Western. As the second season started, the Mouseketeer production numbers shifted more to emphasizing older teen couples. Cubby still appeared in comic bits, but he was more often solo than paired with Karen.