Commons leader Andrea Leadsom has quit the cabinet, saying she no longer believes the government’s approach will deliver Brexit.
Her resignation comes amid a backlash against Theresa May’s Brexit plan from Conservative MPs.
Several cabinet ministers have told the BBC that the PM cannot stay, with one saying it is “the end of the line”.
Mrs Leadsom previously ran for Tory leader but withdrew, clearing the path for Mrs May to become prime minister.
As Commons Leader, she was in charge of organising government business.
Her resignation is the 36th by a minister under Theresa May – 21 of them over Brexit.
In a letter to the prime minister, Mrs Leadsom – MP for South Northamptonshire – said she did not believe “the UK would be a truly sovereign United Kingdom through the deal that is now proposed”.
She also described holding another referendum as “dangerously divisive”, and said she was opposed to the government “willingly facilitating such a concession”.
A second referendum would “risk undermining our Union which is something I passionately want to see strengthened”, she argued.
She also attacked the “breakdown of government processes” saying that Brexit-related legislation proposals had not been “properly scrutinised”.
Back in 2016, at the height of the Tory leadership election, Andrea Leadsom’s name was was one of the best known in Westminster.
She was facing off against Theresa May in the final of the race to become prime minister.
But an interview where she appeared to claim she would be a better premier because she had children spelt her downfall, and Leadsom pulled out of the contest.
She was recruited by the victorious – and unopposed May – as environment secretary, before being bumped to the Leader of the House of Commons role following the 2017 election that lost the Tories their majority.
On a day when no other cabinet ministers have yet jumped, the woman who catapulted May into Number Ten nearly three years ago may be the one who finally forces her out.
Downing Street responds to latest resignation to rock cabinet
“Andrea Leadsom has served with distinction and great ability as a member of the government, and the prime minister is grateful for all of her work,” a Number 10 spokesperson says.
“We are disappointed that she has chosen to resign, and the prime minister remains focused on delivering the Brexit people voted for.”
Labour call for a general election
Ian Lavery, chair of the Labour Party, says:
“The Prime Minister’s authority is shot and her time is up.
“While the Tories are ripping themselves apart, our country is in crisis. The government has made a catastrophic mess of the Brexit negotiations, our steel industry is under threat and universal credit is pushing people into poverty.
“For the sake of the country, Theresa May needs to go, and we need an immediate general election.”