[1] In 1535, despite a papal ban, he married Giulia da Varano, daughter of the duke of Camerino and Caterina Cybo.

Guidobaldo and Giulia had a child, Virginia Varana della Rovere, who married Frederico Borromeo.

[4] In 1546, he received a condotta as military leader (Governatore) by the Republic of Venice, for which his father had been a valiant commander during the Italian Wars.

In 1559, he was hired by the King of Spain, helping Bernardo Sanseverino (who had married Isabella, one of Guidobaldo's daughters) in the war against the Ottoman Turks.

On 1 January 1573, a revolt rose against Guidobaldo in Urbino, due to the excessive tax burden that he was exerting over his state.