She was the elder sister of Charles V, Holy Roman Emperor, and her life was dominated by her role in the international dynastic politics of the period.

Her father was also the son of the reigning Holy Roman Emperor Maximilian I and his deceased consort Mary of Burgundy, while her mother was the daughter of the Catholic monarchs, Ferdinand II of Aragon and Isabella I of Castile.

After the death of her father in September 1506 Eleanor was educated at the court of her aunt Margaret of Austria in Mechelen (in modern Belgium).

When she was a child, Eleanor's relatives tried to marry her to the future King of England, Henry VIII, who was seven years her senior.

Her brother Charles arranged the marriage between Eleanor and the King of Portugal to avoid the possibility of Portuguese assistance for any rebellion in Castile.

The Treaty of Cambrai (1529; called La Paz de las Damas - "The Ladies' Peace") paused the conflict between Francis and Charles.

Eleanor left Spain in the company of her future stepsons, who had been held hostage by her brother.

Eleanor was ignored by Francis, who seldom performed his marital obligations and preferred his lover Anne de Pisseleu d'Heilly.

At the official entrance of Eleanor to Paris, Francis displayed himself openly to the public in a window with Anne for two hours.

She lived with her sister in Jarandilla de la Vera,[citation needed] where they often visited their brother, who retired to the Monastery of Yuste nearby.