Proposed job cuts at train operator ScotRail would threaten the safe running of rail services, according to a transport union.
The Transport Salaried Staffs' Association (TSSA) says the ability to run services on time would also be "completely undermined".
It plans to ballot its members on taking industrial action and has written to transport minister Derek Mackay to seek a meeting about plans to axe controllers and train delay attributors.
The ScotRail Alliance, a formal agreement between Abellio ScotRail and Network Rail, was set up in May to create a stronger relationship between the two.
In a letter to Mr Mackay, TSSA general secretary Manuel Cortes said initial consultations with trade unions on the alliance "made clear" that duplication of jobs related to management and that would mean "no operational jobs would be lost".
But he said it was now evident that four jobs will go within the controllers' grade and five train delay attributors will be lost.
Mr Cortes wrote: "It has now been made clear during consultation that four jobs will go within the controller's grade, which TSSA believes would put at risk the safe running of ScotRail.
"It was admitted during the consultation process by management that the current rosters 'were tight', which in short means that there is no room for any jobs cuts within the control grade without leading to overworking and fatigue amongst the controllers left to operate the railway.
"It has also been revealed during consultation that five train delay attributors (TDAs) would also be lost, which with the four control posts would mean that an approach by the alliance for 'on time railway' would be completely undermined and hard to achieve.
"Is the Scottish Government content in allowing safety critical operational jobs to be lost, which in turn puts immense pressure on the staff left to safely operate ScotRail?
"Is the Scottish Government content with the fact that an 'on time railway' is not happening now and never will do with the loss of the TDAs, who are critical in assuring against future delays?"
He added: "This alliance is not an opportunity to save on staff wages but it seems to have become one and one that TSSA will resist."
A spokeswoman for the ScotRail Alliance said: "The creation of the ScotRail Alliance gives us a chance to bring the two businesses together, cut out duplication and deliver a better service for our customers.
"At the moment, we are engaged in consultation with the trades unions about proposed changes to some of our teams. The whole point of these meetings is for proposals to be discussed, views to be taken on board and, where appropriate, changes made.
"We were surprised to see this letter as this is exactly where we are in the process. In fact, we are meeting the trades unions, including the TSSA, for further talks in the week beginning November 23."
Transport Scotland said it supports a "more integrated approach across the whole industry".
A spokesman added: "We therefore welcome closer collaboration which places the needs of passengers at its heart through the delivery of high-quality, reliable services, and a more efficient management and operation of the rail network in Scotland.
"Our new ScotRail franchise rightly places high demands on the operator, from new trains and improved punctuality of services to improved station facilities and a smart-enabled network. We will look to this innovative partnership to help meet and exceed our ambitious plans."
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