Boris Johnson will face a vote in his leadership, a senior Tory MP announced on Monday.
Sir Graham Brady, chairman of the 1922 committee of backbench Conservative MPs, said he had received at least 54 letters from MPs saying they no longer had confidence in the Prime Minister.
Once the threshold of 54 is reached, it triggers a confidence vote which is set to take place between 6pm and 8pm on Monday.
Shortly after 8am, Sir Graham gave a statement outside Parliament, in which he said: “The threshold of 15 per cent of the parliamentary party seeking a vote of confidence in the leader of the Conservative Party has been exceeded.
“In accordance with the rules, a ballot will be held between 1800 and 2000 today Monday 6th June - details to be confirmed.
“The votes will be counted immediately afterwards. An announcement will be made at a time to be advised. Arrangements for the announcement will be released later today.”
Some MPs had “post-dated” their no confidence letters to take effect after the end of the Platinum Jubilee bank holiday long weekend, Sir Graham confirmed.
Shortly after his statement, Downing Street said Mr Johnson "welcomes the opportunity to make his case to MPs".
A No10 spokeswoman said the vote on Monday night was "a chance to end months of speculation and allow the Government to draw a line and move on".
If Mr Johnson gains a majority in the vote, 180 MPs backing him out of 359, he can continue as Prime Minister.
However, he could struggle to stay in No10 if there is a major revolt of at least 100 Tory MPs, even if he gets a majority.
It comes after Mr Johnson lost the support of a long-time ally on Monday.
Former minister Jesse Norman tweeted: “I have supported Boris Johnson for 15 years, for the London Mayoralty and for PM.
“Very sadly, I have written to him to say I can no longer do so.”
Mr Norman said that Boris Johnson’s current policy priorities were “deeply questionable” and that there were no circumstances in which he could serve in a government led by him.
In his letter to the Prime Minister, Mr Norman, a former minister, warned that any breach of the Northern Irish Protocol would be “economically very damaging, politically foolhardy and almost certainly illegal”.
“You are the leader of the Conservative and Unionist party, yet you are putting the Union itself gravely at risk,” he said.
He said the Government’s Rwanda policy was “ugly, likely to be counterproductive and of doubtful legality” and that plans to privatise Channel 4 were “unnecessary and provocative”.
Earlier, one of Mr Johnson’s key allies warned that moves to topple the Prime Minister were “inexcusable”.
Steve Barclay, the No10 chief-of-staff, led a fightback to defend the PM, arguing that the next election would not be decided by the furore over the Sue Gray report into the partygate scandal.
Mr Barclay highlighted the Prime Minister’s leadership over the Ukraine conflict, that he had got “Brexit done” (despite the economic harm it has done to the country), that the Government had protected jobs during the Covid pandemic, was seeking to level up the country, as well as other reforms.
Writing on the Conservativehome website, he added: “The problems we face aren’t easy to solve. Democracies around the world are all currently facing similar challenges. But under Boris Johnson’s leadership, our plan for jobs shows how we are navigated through these global challenges.
“To disrupt that progress now would be inexcusable to many who lent their vote to us for the first time at the last general election, and who want to see our Prime Minister deliver the changes promised for their communities.”
