Margaret's marriage to the Protestant Henry of Navarre in August 1572 was intended to strengthen a settlement between rival religious and dynastic interests. The St. Bartholomew's Day massacre followed within days. In her later memoirs she recalled sheltering a wounded Protestant in her rooms. During the following years she navigated disputes involving her husband, her brother Henry III, and her younger brother Francis of Alençon.

She joined Henry of Navarre's court in the southwest and participated in factional politics during the French Wars of Religion. Her relationships with both Henrys deteriorated, and an attempt to establish her position at Agen in 1585 failed. Captured the following year, she was confined at Usson in Auvergne. She eventually gained substantial freedom within the castle, maintained a household and circle of visitors, and remained there until 1605. During that period she began writing memoirs in response to Brantôme's account of her life.

Henry of Navarre's accession as Henry IV in 1589 made her queen of France, though the couple remained separated. Their marriage was annulled in 1599 after negotiations. Back in Paris from 1605, she retained the title of queen and supported writers, musicians, religious foundations, and charitable works. She reconciled with Henry and developed ties with his new family. Her memoirs, published after her death, offered an account of court politics from a royal woman's perspective. She died in Paris in March 1615.