London Mayor Boris Johnson has withdrawn his support for the HS2 high-speed rail link between London and the North, calling the project "perverse" and "inadequate".
The intervention from Johnson, the most influential figure to oppose the project, will come as a blow to David Cameron who just days ago insisted the government was committed to the rail link.
Johnson says the whole stretch of the proposed line which falls within London should be routed through tunnels, which would add massively to the £34 billion estimated cost of the new link to Birmingham, Leeds and Manchester.
The Mayor says the project would double the number of passengers arriving at Euston station each morning, and he would not back it without government commitment to increase London Underground capacity.
Mr Johnson's comments come in a letter to anti-HS2 campaigner Jerry Marshall, obtained by a Sunday newspaper, in which he lists his objections to HS2.
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The Mayor writes in his letter, dated June 20: "While I have expressed support for a high speed rail network in the past (and continue to support it in principle) my support is conditional on a number of specific criteria and on the need to make the new railway work well for London.
"The proposal now being consulted on does not reflect these conditions and is inadequate for a number of reasons."
Proposals to send the track along an elevated section at Hillingdon and at ground level through Ealing raised "significant environmental concerns", said Mr Johnson, who faces a mayoral re-election battle against Ken Livingstone next year.
"Elsewhere along the route, particularly outside of London, considerable effort has gone into mitigating noise and visual effects by redesigning the railway alignment to screen it from surrounding areas or by burying it in tunnel," he said.
"It is perverse that a section of the route through Greater London, clearly affecting large numbers of people, has been subject to so little environmental mitigation.
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"I am seeking substantial changes in design of the route to ensure these impacts are properly addressed, preferably by tunnelling the whole route through London. Without such changes I cannot support the current proposal."
Warning the Underground may not be able to cope with the influx of extra passengers if HS2 is built, Johnson said: "I wanted a commitment from the Government that their proposals for HS2 would include new Underground rail capacity between Euston and Victoria. They do not and on this basis I cannot support the current proposal."
If ministers do not meet the Mayor’s demands, and his objections to the project continue after negotiations, it could mean massive delays or even the scrapping of HS2.
HS2 is opposed by local conservative MPs including The Speaker, John Bercow (Buckingham), Welsh Secretary, Cheryl Gillan (Amersham) and Minister for Europe, David Lidington (Aylesbury)
Various anti-HS2 campaigners are raising awareness of the government's controversial plans for the high speed link, which will cut through swathes of areas of outstanding natural beauty. The Government is currently engaged in a formal consultation regarding HS2 which closes on July 29.
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The Mayor, who is widely suspected of positioning himself as a possible successor to Mr Cameron as Conservative leader, has also clashed with the PM over Europe, immigration, tax and cuts to housing benefit.