On the West Coast main line, linking Glasgow Central to London Euston, engineers will renew tracks at several locations including Carstairs, Beattock and Symington South.
As part of this programme to help make the railway infrastructure more resilient and reliable, structural improvement works will also take place at Cathcart Road in Glasgow.
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Most of the country's services are unaffected, but due to the complexity of the engineering work, the programme cannot be delivered without some short-term closures of the line.
Liam Sumpter, Network Rail route director for Scotland, said: "The improvement works planned for the May Bank Holiday are crucial to improving the reliability of Scotland's Railway.
"The work will help to provide a more modern railway while limiting future disruption for passengers.
"We understand the inconvenience this work will cause some customers, but such a significant programme cannot be delivered without temporary disruption.
"We would like to reassure passengers every effort has been made to reduce the impact of our activities as much as possible and we thank you for your understanding."
