He was the subject of two famous portraits by Agnolo Bronzino, one as an infant and another of some years later, together with Eleonora of Toledo (although the subject of the latter has been identified also as Francesco or Garzia).
Probably already suffering from tuberculosis, Giovanni died two years later in Livorno, from a malaria attack. His mother and his brother Garzia died of the same illness a few days later.
Centuries after his death, a myth arose saying that his brother Garzia, killed him, following a dispute in 1562. In turn, his father Cosimo, furious, killed Garzia with his own sword. However, modern exhumations showed no signs of violence on the bodies and found to have died together of malaria in 1562.
His father Cosimo had another son in 1563, who was called with the same name (he is best known as Don Giovanni de' Medici).