Finance firm Standard Life has served notice that it could move some of its operations out of Scotland if the country votes for independence.
The pensions and savings company - which has been based in Scotland for 189 years - said today it would "take whatever action we consider necessary" to give continuity to its customers.
Its annual report revealed it has already begun to set up additional registered companies outside of Scotland "into which we could transfer parts of our operations if it was necessary to do so".
Chief executive David Nish said the action had been taken amid continued uncertainty ahead of the Scottish independence referendum, to be held on September 18 this year.
He said there was still doubt over a "number of material issues" - including what currency an independent Scotland would use and what the arrangements for financial regulation would be.
"We will continue to seek clarity on these matters, but uncertainty is likely to remain," he said.
"In view of this, there are steps we will take based on our analysis of the risks."
The move by Standard Life comes after Chancellor George Osborne and his Labour and Liberal Democrat counterparts all ruled out entering into a formal currency union with an independent Scotland - the Scottish Government's preferred option, which would allow the country to continue to use the pound if it left the UK.
Mr Nish stressed work to establish the new registered companies was a "precautionary measure to ensure continuity of our businesses' competitive position and to protect the interests of our stakeholders" .
He added: "As chief executive, my commitment is whatever happens, we will continue to serve the needs of our customers and maintain our competitive position."
Edinburgh-based Standard Life employs about 5,000 people in Scotland - more than half of its total workforce of 8,500.
Deputy Prime Minister Nick Clegg said the move did not surprise him, because of the uncertainty over issues such as an independent Scotland's currency and membership of the EU.
He told BBC Radio 5 Live: "Because of the failure of the SNP to prepare for this moment and spell out what they mean by independence, it is no wonder that major employers are saying 'Maybe we can't continue with our presence north of the border'."
But Scottish Finance Secretary John Swinney said he looked forward to Standard Life "continuing to play its part in building that strong Scottish economy in the future".
He said: "When Standard Life previously expressed concerns about the consequences of devolution, these concerns ultimately proved to be unfounded and the company has successfully continued to grow its business here, underlined by the announcement just this month of a £75 million acquisition in central Edinburgh described as a 'long-term investment'.
"Standard Life's strengths lie in its workforce here in Scotland. We are very happy to engage with the company to address the issues raised in their annual report and we look forward to the company continuing to play its part in building that strong Scottish economy in the future.
" Standard Life's comments show exactly why our proposals for a formal currency area are the right proposals, why they are in the best interests of business on both sides of the border and why that is what will be implemented by both governments."
The Scottish Government believes UK ministers will adopt a different approach to a currency union if there is a Yes vote in the referendum, and Mr Swinney said: " The UK Government are engaged in a systematic campaign of bluff, bluster and bullying, but we have already seen the UK Government accept that it will remain legally liable for all UK debt, and the Prime Minister offer his support for Scotland's membership of the EU - the issue of currency will be no different.
"In terms of Scotland's position in Europe, an independent Scotland will remain in the EU, and the biggest threat to that membership comes from Westminster's proposed in-out referendum.
"The way to provide clarity and certainty is through common sense, mature discussions which will take place in the 18 months following a Yes vote - and technical discussions could and should take place now."
Blair Jenkins, chief executive of the pro-independence campaign Yes Scotland, said: " Standard Life wants to see agreements on currency, regulation and taxation, which is exactly what the Scottish Government has proposed.
"Standard Life wants a formal currency union and so do we. The only threat to that comes from the refusal of the No campaign and the UK Government to get involved in sensible discussions.
"We know common sense will win the day, just as David Cameron has agreed that the UK would support Scotland as an independent member of the EU, in the same way there will be a sterling area after independence because it is clearly in the best interests of both Scotland and the rest of the UK and that means companies like Standard Life will continue to flourish and operate fully in Scotland."
The Royal Bank of Scotland, which also reported annual results today, said it was ''politically neutral'' over Scottish independence, but warned a vote in favour could impact the group by affecting its credit rating.
It said: ''The group's borrowing costs and its access to the debt capital markets and other sources of liquidity depend significantly on its and the UK Government's credit ratings, which would be likely to be negatively impacted by political events, such as an affirmative outcome of the referendum for the independence of Scotland.''
The bank stressed it was ''impossible'' to quantify the financial hit of a Yes vote.
It added: ''Clearly there are issues we are looking at - currency, the application of financial regulation, lender of last resort, credit ratings - which could affect us.
''But there is real uncertainty about how any of these matters would be settled in the event of a Yes vote and the outcome would depend on negotiations between the two governments.''
Chief Secretary to the Treasury Danny Alexander said the positions of Standard Life and RBS showed "the risks of independence becoming ever clearer".
He said: "It's common-sense that when you have something that works there will be adverse consequences if you rip it apart.
"The strength and stability of the United Kingdom is the essential underpinning of Scotland's successful financial services sector over several centuries.
"These businesses are reasonably and fairly setting out the consequences of the SNP's dangerous, risky, and unclear plans for independence. I doubt they'll be the last."
Former chancellor and leader of the pro-union Better Together campaign, Alistair Darling, said an "obsession" with independence was putting thousands of Scottish jobs at risk.
He said: "Independence will cost jobs. This is the reality of Scotland leaving the UK and losing the UK pound. Companies like Standard life rely on the strength, security and stability of the UK.
"Scotland's financial services industry employs 200,000 people but these jobs and the standard of living of Scots are under threat because of Alex Salmond's obsession with independence.
"As both Standard Life and RBS make clear today, Alex Salmond's complete failure to set out a credible position on currency puts jobs in Scotland at risk. A currency union is not going to happen, so we need to know what money our economy would be using."
Liz Cameron, chief executive of Scottish Chambers of Commerce, said: " Businesses can plan best in a stable economic environment.
"The referendum is necessarily creating some degree of uncertainty and businesses will naturally work with this and plan accordingly, so whether Scotland chooses to vote 'yes' or 'no', they will need stronger detail from both sides of the campaign on issues such as tax, currency and European Union membership in order that they can best plan for the future."