The 60-hour track closure from early Saturday to 6pm Monday will involve more than 200 workers carrying out more than 12 projects in the $235 million upgrade.

The majority of works will take place south of Benalla.

It marks the first "possession" period for major contractor John Holland. which has fully established a project office in Wangaratta where more than 100 people are working full-time.

"A key focus of the North-East rail line upgrade is to ensure regional centres in North-East Victoria directly benefit and more than 18 local businesses are already contracted to work on the multi-million-dollar project," Australian Rail Track Corporation's Victorian projects general manager Ed Walker said.

"Matt Tennant and his team from Seymour-based contractor, TENEX Rail, have been working on the project since early 2019.

"With a major pipeline of project work booked, they have been able to invest almost $1 million in brand new railway maintenance machines, upskill and extend the hours of existing staff as well as create several additional positions for locals."

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Mr Walker has requested the public to take extra care around worksites as a greater number of trucks and machinery will be in operation.

There may also be some road closures and detours to be familiar with due to level crossing renewal works.

"While every effort will be made to minimise disruptions - we would like to thank the community for its patience with us while these essential works are being delivered," Mr Walker said.

Level crossing upgrades are taking renewals at two locations in Heathcote Junction and also at Tallarook and Seymour.

Bridge deck replacements will also take place at locations south of Seymour with ARTC also delivering track and level crossing works closer to Melbourne.

Works beginning early last year included ballast and track improvement, timber bridges upgrades, level crossing renewals and installation of new rail crossovers before major contract works started last month from Wodonga.

A major ballast stockpile has also been established at Violet Town and the ARTC is also progressing a $14 million signalling improvement project in Melbourne.

The line recently shut for more than a week due to the Wallan derailment.