Zahran Alloush, head of the powerful Jaish al-Islam Syrian rebel group, was killed on Friday east of Damascus, a monitoring group and Syria's opposition said.
His death "stands as one of the most significant opposition losses" of Syria's nearly five-year uprising, analyst Charles Lister said on Twitter.
The Syrian Observatory for Human Rights monitor said Alloush and five other commanders were killed "in an air strike that targeted one of their meetings in Eastern Ghouta."
The death of Alloush, 44, was confirmed on Twitter by the head of Syria's opposition National Coalition, who posted a message of condolence.
Syria's state television channel also reported the death by air strike, but did not say who had carried out the raid.
Jaish al-Islam is the most prominent rebel faction in the Eastern Ghouta region, an opposition bastion east of the capital, and was recently represented at landmark opposition talks in Saudi Arabia.
The Syrian government regularly refers to the group as "terrorists".
Alloush's death comes after Syria's army announced a massive operation to retake rebel-held Eastern Ghouta. The regime's forces have been backed by Russian air strikes since September 30.
Jaish al-Islam was known to have extremist views and to have supported the establishment of an Islamic state before recently moving towards a more moderate position.
Alloush spent at least two years in Syrian prisons before being released in a general amnesty in June 2011.