The fire broke out around 10 a.m. Monday at the Eugene O'Neill Theatre on West 49th Street.
The Broadway venue is currently showing "The Book of Mormon," but no performances are scheduled for Monday.
Firefighters could be seen fighting the blaze from the roof as smoke rose into the air.
FDNY Assistant Chief David Simms said that the fire broke out originally in the electrical room, between the 4th and 5th floor.
The fire department apparently worked to knock down the original fire and then discovered more fire between the 5th floor and the roof.
The FDNY said this was a "deep seeded fire" involving electrical equipment and overall was very difficult to get to.
The theater is out of service pending repairs. The Department of Buildings is on the scene and will investigate.
There is substantial damage on the fourth-floor electrical room which contains lighting equipment and hanging chandeliers.
One firefighter suffered minor injuries and all theater personnel are accounted for.
The cause of the fire is under investigation.
The Ambassador Theater Group, which represents "The Book of Mormon" released a statement saying:
"The FDNY is at the site of the fire now and are handling matters at the Eugene O'Neill Theatre. Everyone has been safely removed from the theater. We will keep all parties abreast of this developing situation."
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