The son of French nobility, Aubusson joined the Knights of St (Hospitallers). John c. 1453. The Knights, with their headquarters at Rhodes, held the island as a bar to Ottoman expansion in the Aegean. Aubusson became grand master of the order in 1476 and in 1480 gained widespread fame in Europe for successfully defending Rhodes against Sultan Mehmed II's fleet.

Later, when Cem (Jem), brother of Mehmed's successor Bayezid II, took refuge in Rhodes, Aubusson, despite his promise to Cem of safe conduct, accepted a bribe from Bayezid and had Cem imprisoned. After six years, Aubusson turned Cem over to Pope Innocent VIII, who had been vying with the kings of Hungary and Naples for the possession of so valuable a political weapon as the Sultan's brother. In return, Innocent made Aubusson a cardinal (1489) with the power to confer all benefices connected with the order without the sanction of the papacy. As cardinal, Aubusson reformed the Order of St. John, strengthened its authority in Rhodes, and eliminated Judaism from the island by expelling all adult Jews and forcibly baptizing their children.

Two years before his death, Aubusson failed in trying to organize a large international crusade against the Turks.