Eddie Izzard has chosen to use the pronouns "she" and "her", and says she wants "to be based in girl mode from now on".
The gender-fluid star said it "feels great" and it is the first time she has asked to be referred to using the pronouns while making a television programme.
Izzard, 58, has previously identified as a transvestite, frequently wearing dresses, make-up, heels and jewellery both on and off stage.
She has also called herself "a lesbian trapped in a man's body", and said she feels like "a complete boy plus half a girl".
LGBT charity Stonewall praised the comedian for her bravery and said her openness would have an impact on transgender communities worldwide.
Associate director of communications and campaigns Robbie de Santos said: "Being open about using different pronouns is often incredibly difficult, especially in the public eye – her courageous decision will mean a lot to trans and gender-fluid communities around the world."
Comedian Shappi Khorsandi also praised Izzard, tweeting on Sunday: "I see Eddie Izzard is trending. I can't tell you what she means to me as a comic.
"Rocked my comedy world when I was a teen and beyond. Changed everything, made room. I love her and this morning I'm very happy for her."
In the past, Izzard has expressed the view that being transgender is caused by genetics.
Meanwhile actor Elliot Page, who came out as transgender earlier this month, thanked fans for their "love and support".
The 33-year-old Umbrella Academy star, who has adopted the pronouns he and they, told fans it felt "remarkable" to finally pursue his "authentic self".
Away from the positive message, Page also highlighted the high levels of violence towards transgender people and the 40% rate of attempted suicide among the trans adult community.