Multiple episodes center LGBTQ+ storylines, including Kurt Hummel's intensified bullying arc and his meeting with openly gay Blaine Anderson in "Never Been Kissed," Santana Lopez's closeted struggles and prom queen scheme involving blackmailing bully Dave Karofsky in "Born This Way," and the development of same-sex pairings like Klaine and Brittana. The "Born This Way" episode explicitly structures plots around homosexuality, self-acceptance via Lady Gaga's title song, and characters displaying personal "flaws" on T-shirts, with Santana's "Lebanese" shirt underscoring lesbian identity as a focal point. Creator Ryan Murphy has stated his intent to use the series for personal visibility of gay characters and to foster acceptance by showing nontraditional families and experiences to mainstream audiences.

This drives character arcs and subplots beyond organic storytelling, with explicit messaging on homophobia, bullying tied to sexuality, and embracing "otherness." Racial and disability diversity appears in the cast (e.g., Artie in a wheelchair, characters like Mercedes and Tina), but the season's appeal and narrative weight lean heavily on identity politics framing rather than incidental inclusion. Audience reception included praise for representation alongside criticism of heavy-handed preachiness, with conservative pushback on the homosexuality depictions.