Joo Jin-mo debuted as an actor in the 1999 movie Dance Dance. That same year, he acted alongside Choi Min-shik and Jeon Do-yeon in the film Happy End, which won him Best Supporting Actor at the 37th Grand Bell Awards. In 2000, he starred in the KBS2 drama Look Back in Anger, which earned him the Best New Actor trophy at the KBS Drama Awards. In 2003, he took home an Excellence Award at the SBS Drama Awards for his work in Punch. In 2006, he led the widely popular musical romantic comedy film 200 Pound Beauty along with Kim Ah-joong. In 2008, he played a homosexual Goryeo king in the erotic period film A Frozen Flower, which gave him his first Best Actor trophy at the 45th Baeksang Arts Awards. In 2013, he joined the main cast of the hit MBC sageuk Empress Ki, which scored him a Top Excellence Award at the MBC Drama Awards.

Joo Jin-mo and medical doctor Min Hye-yeon tied the knot in June 2019 on Jeju Island. In 2020, the actor’s personal text messages containing photos of women and vulgar comments about them were exposed by phone hackers. He has since apologized to the women mentioned in the text messages.