The Other Boleyn Girl follows the lives of the Boleyn sisters, Mary and Anne, beginning before Anne became Queen of England by marriage to self-indulgent King Henry VIII. Before he began his courtship with Anne, the King favored Mary, at the time married to her first husband, William Carey. Mary Boleyn was a lady-in-waiting to the monarch's first wife, Catherine of Aragon. Despite her objections, Mary is forces into becoming one of the King's mistresses with her husband's reluctant, yet mandatory, agreement and permission. Mary asks her husband if she has offended him and he claims she has not, but Mary is troubled that he did not object strongly enough to her illicit relationship with the King. Mary is weighted down by feelings of guilt whenever she serves Queen Catherine as the Wueen is aware if her affair yet says nothing. Soon, however, Mary comes to terms with her relationship with the King and begins to fall in live with him.
Meanwhile, Mary's sister Anne falls in love with Lord Henry Percy, a member of the aristocracy and heir to the title of Duke of Northumberland. Lord Henry is already betrothed but Anne is oblivious to this and the two act flirtatiously when around each other. Despite Mary's admonishment they soon consummate their affair whilst Mary is at the pinnacle of her power as the King's primary mistress. Courageously, Mary shares her concerns with their parents and their uncle who all condemn Anne's behavior, warning her that she is making a grave mistake in continuing her relationship with a man who is already engaged to be married to the daughter of an Earl - a union that was orchestrated by Cardinal Thomas Wolsey, with the King's permission and blessing. Fearing that Anne's behavior will endanger Mary's relationship with the King, her family plans to send her away. Anne is devastated and will not listen to Mary's explanation and screams at Mary that she will neither forgive nor forget what she has done. Anne leaves in exile to the family seat of Hever Castle.
Mary confesses that she is falling in live with the king and is increasingly enjoying the time that they spend together; she also feels no remorse for Anne's banishment. At Hever, Anne misses Henry Percy and believes she will never fall in love again. She devises a plan that will enable her to be returned to court and feigns compliance. Back at court, Mary is readying herself for a tryst with the King when her husband arrives unexpectedly, asking her to spend the night with him. Mary explains that she cannot, reminding him gently of her duty to the king. He asks her if she will come to him the next night but again she turns him down and he leaves in anger to find another woman to spend his nights with, leaving Mary curiously devastated.
A year goes by and Mary becomes pregnant with a child that is widely believed to be the King's due to her separation from William. Anne returns to court, dutifully telling her family she is honored to be there and expressing her desire to serve the family any way she can. Mary is extremely skeptical and doubts the sincerity of her sister's words. A few days later the family devises a plan for Anne to distract the king as Mary enters her confinement, George Boleyn arranging the introduction whilst the king is walking with other ladies at court. Anne interrupts their conversation and informs him that Mary is glowing and looks lively, but the King takes an interest in Anne, telling her he finds her more attractive than Mary.
Mary gives birth to a son, naming him Henry. The king cares only for Anne and Mary soon leaves court to be with her son, leaving Anne to take her place as his mistress. Mary returns to her husband and they have a daughter together but Mary is distressed when William seems to care little for his child. Two years later William dies from sweating sickness whilst Anne receives new chambers at court. She tells Mary that she needs her by her side at all times and that she must be protected when the king makes their relationships more official. Mary is shocked by this and reminds Anne of her love for Henry Percy but Anne claims to have felt nothing for him, threatening her sister if she ever reveals their relationship.
The King learns of Henry Percy's wife's request for a divorce and informs Anne if this, telling her that they cannot be together if there was ever any relationship between her and Percy. Anne forces Mary to lie and say that no betrothal ever existed; to appease the king, Anne sleeps with him for the first time, prompting him to banish Catherine of Aragon to a remote castle and marrying Anne who quickly becomes pregnant. Anne gives birth to a daughter, not the male heir the King is focused upon, and her hold on him begins to slip away, as he takes a mistress.
Unbeknownst to Anne, Mary slips away from court to meet with William Stafford, a former servant if the Boleyn family who had previously declared his love for her. Mary accepts his proposal of marriage and on hearing this Anne banishes her sister from court, telling her that she had brought shame and disgrace onto the family.
Two years pass; Anne has two miscarriages and cannot seem to produce a son. Feeling increasingly threatened her behavior becomes erratic and in front of all the court she demands to know the identity of the king's mistress. Her rudeness angers the king who drives her to depart from the room, followed by her father who berates her for her behavior. Anne demands respect but her father informs her she now has many enemies.
Thomas Cromwell suggests to the king that he leaves court and journeys to Wulfhall in Wiltshire, home of the Seymour family. The king's time there is spent predominantly with the eldest Seymour daughter, Jane. Back at court Anne finds a sketch of herself being decapitated. She seeks out Feirge and begs him to bring Mary to court. Mary devises a plan that suggests Anne and George lay together in an effort to make Anne pregnant; despite George's protests they go through with the dark actions and Anne indeed becomes pregnant. When she reveals her pregnancy it delights the king but another miscarriage ensures that Anne's days in court are coming to an end. Anne reminds the king that he loved her once and proclaims herself innocent of adultery and incest, but she is led away by guards and two days later is beheaded. Mary leads a happy life of a nobody, blissfully caring for all of the children with her beloved husband William Stafford.