Joanna died aged 75 in 1555, at which point her son Charles, the Holy Roman Emperor, became the sole ruler of Castile and Aragon.

[4] Joanna's academic education consisted of canon and civil law, genealogy and heraldry, grammar, history, languages, mathematics, philosophy, reading, spelling and writing.

Joanna's royal education included court etiquette, dancing, drawing, equestrian skills, music, and the needle arts of embroidery, needlepoint, and sewing.

[5] Joanna entered a proxy marriage at the Palacio de los Vivero in the city of Valladolid, Castile, where her parents had secretly married in 1469.

Joanna began her journey to Brabant in the Low Countries, which consisted of parts of the present day Netherlands, Belgium, Luxembourg, France, and Germany, on 22 August 1496.

"[8]: 315  In response, Ferdinand embarked upon a pro-French policy, marrying Germaine de Foix, niece of Louis XII of France (and his own great-niece), in the hope that she would produce a son to inherit Aragon and perhaps Castile.

Maximilian tried to comfort Joanna with festivities and she spent weeks accompanying him in public events, during which she acted like a wise, prudent queen, as noted by the Venetian ambassador.

When Philip tried to gain support from Castilian nobles and prelates against Ferdinand though, Joanna firmly refused to act against her father.

To the general surprise, Ferdinand had unexpectedly handed over the government of Castile to his "most beloved children", promising to retire to Aragon.

Philip and Ferdinand then signed the Treaty of Villafáfila secretly, agreeing that Joanna's "infirmities and sufferings" made her incapable of ruling and promising to exclude her from government and deprive the Queen of crown and freedom.

Ferdinand promptly repudiated the second agreement the same afternoon, declaring that Joanna should never be deprived of her rights as Queen Proprietress of Castile.

On 12 July,[7]: 69–91  they swore allegiance to Philip I and Joanna together as King and Queen of Castile and León and to their son Charles as their heir-apparent.

A regency council under Archbishop Cisneros was set up, against the queen's orders, but it was unable to manage the growing public disorder; plague and famine devastated the kingdom with supposedly half the population perishing of one or the other.

His arrival coincided with a remission of the plague and famine, a development which quieted the instability and left an impression that his return had restored the health of the kingdom.

On 17 August 1507, three members of the royal council were summoned – supposedly in her name – and ordered to inform the grandees of her father Ferdinand II's return to power: "That they should go to receive his highness and serve him as they would her person and more."

However, she made it evident that this was against her will, by refusing to sign the instructions and issuing a statement that as queen regnant she did not endorse the surrender of her own royal powers.

He had Joanna confined in the Royal Palace in Tordesillas, near Valladolid in Castile, in February 1509 after having dismissed all of her faithful servants and having appointed a small retinue accountable to him alone.

[19] The Castilian Cortes, meeting in Valladolid, spited Charles by addressing him only as Su Alteza ("Your Highness") and reserving Majestad ("Majesty") for Joanna.

The kingdoms of Castile and Aragon (and Navarre) remained in personal union until their jurisdictional unification in the early 18th century by the Nueva Planta decrees, while Charles eventually abdicated as Emperor of the Holy Roman Empire in favour of his brother Ferdinand, and as King of Spain in favour of his son Philip – an act that represented the "transition from a universal empire to defence of the interests of the 'Austrian family' (austriacismo), in other words, to a close alliance between two parts of the dynasty, aimed at guaranteeing the hegemony of Catholicism and of the dynasty within Europe".

In an attempt to prevent this, Don Antonio de Rojas Manrique, Bishop of Mallorca, led a delegation of royal councillors to Tordesillas, asking Joanna to sign a document denouncing the Comuneros.

Charles ensured his domination and throne by having his mother confined for the rest of her life in the now-demolished Royal Palace in Tordesillas, Castile.

[9] She is entombed in the Royal Chapel of Granada in Spain, alongside her parents, Isabella I and Ferdinand II, her husband Philip I and her nephew Miguel da Paz, Prince of Asturias.

[27]: 9  She may also have been unjustly painted as "mad" as her husband Philip the Handsome and her father, Ferdinand, had a great deal to gain from Joanna being declared sick or incompetent to rule.

[27]: 12 Bethany Aram argues that while she seemed to be unable or unwilling to rule herself, Joanna's major (political) significance lay with her defense of the rights of her descendants and thus the Habsburg dynasty.

Nalle opines that overall, Joanna was a troubled individual who, not trained for the political world, found herself surrounded by strong personalities, and had to face a shocking amount of cruelty and deceit.