Turkey says Syria fired on one of its planes that was taking part in a rescue operation for a warplane shot down by Syrian forces last Friday.
Turkey’s deputy PM said the CASA search and rescue plane, looking for the F-4 Phantom jet, was not brought down.
He vowed Syria would “not go unpunished” but that Turkey had “no intention” of going to war.
Nato will discuss the downing of the jet on Tuesday at a meeting called by Turkey, a member state.
Speaking at a televised news conference, Turkish Deputy Prime Minister Bulent Arinc did not specify when the second incident took place and did not say whether the search and rescue plane was hit.
He said the Syrians had stopped firing following a warning from the Turkish side.
Mr Arinc said Turkey would protect itself within the framework of international law, but had “no intention of going to war with anyone”.
Syria has insisted the F-4 was engaged while it was inside its airspace.
Syrian foreign ministry spokesman Jihad Makdissi said the location of the wreckage would prove its case.
He said on state TV: “The Syrian defence forces used an anti-aircraft gun with the longest range of 1.2km. We can confirm the damage was caused by anti-aircraft fire. We didn’t use radar for this action.”
But Mr Arinc said: “There is no doubt that the Syrians intentionally shot down our plane in international airspace. The facts in our possession show that our plane was hit by a heat-seeking laser-guided missile.”
He added: “To target an aircraft in this fashion without any warning is a hostile act of the highest order.”
The search is continuing for the jet and its crew, but hopes of the two men being found alive are fading.
Nato members have condemned Syria’s actions and convened a meeting under article 4 of its constitution, which states any member can request talks if it feels its territorial integrity has been threatened.
Article 5 states an attack against any Nato country shall be considered an attack against them all, although diplomats say this is unlikely to be invoked and that the meeting will probably only issue further condemnations.
On Monday, EU foreign policy chief Catherine Ashton urged Turkey to be “restrained in its response”.
German Foreign Minister Guido Westerwelle also urged calm, saying: “De-escalation is crucial at this moment.”
Trickle of defections
Earlier on Monday, Turkish media reported that several high-ranking Syrian military figures had defected to Turkey.
A general, two colonels, two majors and about 30 other soldiers were said to have crossed into Hatay province on Sunday night.
They were part of a group of some 200 people who crossed the border, the Anatolia news agency said.
There has been a steady trickle of defections from the Syrian armed forces over the past year, most of them to opposition forces fighting inside the country.
The BBC’s Jonathan Head in Istanbul says this is one of the biggest single groups of soldiers to defect to Turkey but so far there is no evidence that they have had a significant impact on the Syrian military’s ability to fight.