Theresa May has been urged by the House of Lords Speaker not to stuff the upper chamber with Tory peers or threaten it with abolition to ensure her Brexit plans get through Parliament.
Lord Fowler said the Lords would not "sabotage" Brexit, although he insisted peers must keep their role in "improving" legislation from the Commons.
His intervention comes as the Prime Minister awaits a Supreme Court ruling expected this month on whether she must get Parliament's approval before triggering the formal Brexit process under Article 50 of the EU treaties.
If the court rules against the Government, Mrs May expects legislation to approve the triggering of Article 50 to pass through the Commons, given Labour's commitment not to block or delay Brexit.
But there have been suggestions that the PM's plan to begin the process by April may be held up in the Lords, where the Tories do not have a majority, by anti-Brexit peers.
Lord Fowler's insistence that peers should be able to "improve" legislation could still spark concerns that the Lords will seek to amend any Article 50 Bill and therefore potentially hold up its passage through Parliament.
Writing in the Telegraph, the Lords Speaker said: "The Lords recognise the primacy of the Commons based on the fact that they are the elected chamber and we are not.
"In return most MPs value the check that scrutiny by the Lords provides. We are not here to sabotage legislation – we are here to improve it."
Cabinet ministers have reportedly told Mrs May to threaten to abolish the Lords if peers seek to frustrate Brexit but the Lords Speaker warned her against it.
He wrote: "Some politicians seem intent on creating discord between the House of Commons and the House of Lords.
"Although we do not yet know the decision of the Supreme Court on triggering Article 50, let alone the contents of the Great Repeal Bill, a number of MPs and indeed ministers have made threats aimed at the very existence of the Lords if there is any delay in the Brexit process.
"One Cabinet minister has been quoted (anonymously of course) as saying that any disagreement between Lords and Commons should be followed by a one line bill called 'the abolition of the Lords'.
"Another MP mused on the radio that if there was disagreement, the Prime Minister might create over 100 new Peers to ensure the Government gets its way.
"It is not my role to enter into the politics of Brexit, but I do think that I am entitled to defend the House of Lords from this kind of attack."
Meanwhile, 30 leading British entrepreneurs urged the PM to keep the UK in the European single market and avoid a swift reduction in immigration after Brexit.
In a letter to the Financial Times, the founders of companies such as Jack Wills, eBookers, Games Workshop and Zoopla urged Mrs May to "protect the economy of the future" and keep Britain "open to trade and talent".
The call, organised by Open Britain, comes as the Prime Minister prepares to set out her broad approach to leaving the European Union in a speech later this month.
She has made immigration controls a top priority for Brexit negotiations, which suggests Britain will leave the single market because EU leaders insisted free movement of its citizens is a condition of membership.
Elsewhere, the Department for Exiting the EU (DExEU) strongly denied reports that its permanent secretary, Olly Robbins, opposed the appointment of Sir Tim Barrow as the UK's new ambassador to the EU and sought to downgrade the role so the department could take control of negotiations in Brussels.
A DExEU spokeswoman described the claims as "fundamentally untrue", adding: "Sir Tim will play a crucial role in the negotiation, guiding UKRep throughout the negotiation period and helping the UK forge a positive new relationship with the EU.
"The Department for Exiting the EU is fully behind Sir Tim's appointment and we are looking forward to working with him closely."
It comes after the surprise resignation of Sir Ivan Rogers as ambassador to the EU exposed concerns about the Government's state of readiness for withdrawal negotiations.
Several former top Whitehall officials have warned that the Government has lost a Europe expert at a vital moment while MPs opposed to a hard Brexit said his resignation showed the Government lacks a Brexit plan.