In the film, an ageing lesbian television actress, June "George" Buckridge (Beryl Reid, reprising her role from the stage play), simultaneously faces the loss of her popular television role and the breakdown of her long-term relationship with a younger woman (Susannah York).

[8] Middle-aged actress June Buckridge (Beryl Reid) plays "Sister George", a lovable motor scooter-riding district nurse and local sage, in a fictional long-running BBC television soap opera, Applehurst.

Her worry affects her already volatile relationship with her live-in lover, a younger, beautiful woman named Alice (Susannah York), whom June calls "Childie".

Childlike Alice plays with dolls, writes poetry, and has a minor fashion industry job, but relies on June for most of her financial and emotional support.

The nuns' Mother Superior and the Archbishop complain to the BBC, causing powerful network producer Mrs Croft (Coral Browne) to visit June at home and lecture her about her behaviour and her attitude at work.

Mrs Croft further infuriates June by taking an interest in Alice, complimenting her cooking and encouraging her poetic aspirations.

June's spirits improve when she receives the next script, showing that Sister George has recovered and returned to riding her scooter.

June, upset, tries to sabotage her fellow actors' performances and to drag out the filming as long as possible; afterward, she finds out that Alice lied to her in order to meet with Mrs Croft.

At her farewell cast party, June makes a scene, insulting the senior network executive, pouring drinks over a fellow cast member, and finally exploding in front of the guests when Mrs Croft offers her a new role as the voice of a talking cow on a children's puppet show.

Left alone, June wanders onto the deserted Applehurst set, destroying equipment and props and uttering "moo" like a cow.

"[11] In the movie, Applehurst was changed from a radio programme to a television soap opera, and the lesbian themes of the story were made more explicit, most notably by adding the sex scene between Alice and Mrs. Croft that is not in the stage play.

Browne revealed that Susannah York was deeply upset about the scene and was eating grapes in between takes to stop herself from being physically sick.

Largely on the basis of the graphic sex scene between Alice and Mrs. Croft, The Killing of Sister George received an X rating, which limited its screening in US cinemas and ability to be advertised in mainstream newspapers.

Rarely seen on American television, it was broadcast uncut by Turner Classic Movies as part of a June 2007 salute to gay cinema.

Renata Adler of The New York Times was critical of the film, not just for displaying lesbians, but also for actress performances and the camera work.

[23] On 27 August 2018, it was announced that Kino Lorber would be releasing The Killing of Sister George on Blu-ray in the United States and Canada.