The meeting is the second time key players Moscow, Ankara and Tehran have brought the warring sides together, and comes ahead of a new round of UN-led talks on Syria in Geneva on February 23.
The latest round of discussions is expected to focus mostly on bolstering a faltering six-week truce and has been billed as a prelude to broader negotiations in Geneva.
The regional brokers met separately for talks with regime and rebel delegates today in the Kazakh capital Astana.
A first set of talks on Syria that took place in Astana in January saw the rebels refuse to talk directly to the regime and did not result in any significant breakthrough.
Rebel spokesman Yehya al-Aridi told AFP Wednesday the opposition was sending a a "smaller" delegation than the one it sent for the talks last month.
The rebels, who initially cast doubt over their participation in the latest meeting, will be led by Mohammad Alloush, a leading figure of the Army of Islam (Jaish al-Islam), he confirmed.
Moscow has increasingly taken the lead on pushing talks over Syria after its military intervention on the side of leader Bashar al-Assad helped turn the tables in the protracted conflict.
While Russia and Iran back the regime with military might, Turkey has supported rebels fighting to oust Assad.
Moscow says the Astana process is meant to support the Geneva talks, but there has been speculation that it is working with Ankara to cut the West out of shaping Syria's future.
As the talks were set to begin in Astana, UN envoy Staffan de Mistura was in Moscow for talks with Foreign Minister Sergei Lavrov and Defence Minister Sergei Shoigu.
De Mistura's office is being represented by a technical team at the Astana meet.
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