Harold was born in Decatur, Georgia, the son of an engineer father and a mother who was a real estate agent. The second of three children, Harold's parents were devout Pentecostals, and Harold had a strict Pentecostal upbringing. At age 15, he left the church, saying that he "knew it was bullshit." Harold's father left the church several years later.

After graduating from The Lovett School in Atlanta, Georgia, Harold attended American University in Washington, DC, on a soccer scholarship. He began a Liberal Arts degree in romance literature, only to depart after a year and a half following a conflict with his coach. Harold then moved to San Francisco, California, United States to pursue an interest in photography at the San Francisco Art Institute. He worked a variety of jobs including positions as a Ducati motorcycle technician and a construction worker.

In 1997, friend Susan Landau, daughter of actor Martin Landau, suggested Harold try acting. He relocated to Los Angeles and began a 3-year period of intensive drama study. At 28, he was accepted into the Actors Conservatory Program with the classical theater company A Noise Within. In his theatrical debut, Harold appeared as "Bunny" in Me and My Friends. In 2003, he starred in Wake, produced by Susan Landau Finch and directed by her husband Henry Leroy Finch. The movie featured a cameo by Martin Landau and the lead part of Kyle Riven was written specifically for Harold.

In 2000, Harold landed the controversial role of unapologetic gay man Brian Kinney, a central character on Showtime's popular gay drama Queer as Folk, a breakthrough performance that included a level of explicit gay male sex unusual for American television. Brian Kinney's character, as well as the show itself, elicited quite a great deal of controversy. It was alternately lauded and criticized for its explicit depictions of gay club life. The show ran for five successful seasons, ending in 2005.

Harold had the lead role of Special Agent Graham Kelton in the short lived FOX series Vanished in 2006, but his character was killed off in the seventh episode and appeared only as a corpse in the eighth episode.

Harold also guest-starred as Wyatt Earp in two episodes of the HBO series Deadwood and appeared twice on the CBS series The Unit. Alongside childhood idol David Bowie, Gale is an associate producer of the documentary Scott Walker: 30 Century Man.

Gale Harold returned to the New York stage in Tennessee Williams' play Suddenly Last Summer on November 15, 2006, in the role of Dr. Cukrowicz ("Dr. Sugar"). Harold's co-stars in this Roundabout Theatre repertory production, a limited Off-Broadway engagement running through January 20, 2007, were Blythe Danner and Carla Gugino.

Harold plays the male lead in the indie romantic comedy Falling for Grace, which debuted favorably at the 2006 Tribeca Film Festival (under working title East Broadway). Harold plays an eligible New York bachelor in an inter-racial relationship with an Asian-American woman. The film has been screened at theaters and campuses nationwide and will be available on DVD in March, 2010.

Gale appeared in November 2007 in a guest role on ABC's Grey's Anatomy as Shane, a paramedic and white supremacist with a swastika tattooed on his abdomen, who is injured in an ambulance crash.

Harold joined the cast of Desperate Housewives on the fourth season finale May 18, 2008 as Jackson Braddock, Susan Mayer's love interest. Six months after his serious motorcycle accident, Harold returned to play Jackson on the May 3, 2009 episode of the show.