Local authorities and partner organisations can apply for "capital investment for projects that promote economic regeneration and serve the aims of wider sustainable development", the government said in a statement.
Activities should be "focused on the individuals and areas most in need", it added. The aim is to "help create jobs, enhance skills and employability, and create the environment for businesses to grow and thrive".
According to government guidance on how to apply, a total budget of up to £100 million is available over an initial three-year phase, from April 2018 to 2021.
A national regeneration investment panel will be set up to consider applications and make recommendations to ministers, making sure "that the investment available is utilised as effectively as possible across Wales".
The investment will be "targeted in a small number of regeneration areas in order to generate a greater impact, rather than spreading resources thinly in all parts of Wales", the guidance adds.
According to the document, regeneration investments will be invited "to come forward on a rolling basis", with no deadline for applications.
The guidance asks local authorities to work together at a regional level to identify priority projects. It says: "Our intention is that this programme will be embedded into a new regionally focused model of economic development and regeneration."
Launching the fund on Friday, Welsh cabinet secretary for communities and children Carl Sargeant said the programme "has a crucial part to play in driving prosperity and building resilient communities in all parts of Wales, rather than just in those areas that offer the best commercial returns".
Sargeant said: "Local authorities and regional partnerships should use these funds to complement and reinforce the other investments we are making to widen prosperity, such as the work being pursued under the city deals, our investment in the metros, proposals from the Valleys Taskforce and the work to prepare for Wylfa Newydd."
The guidance on applying for funding can be found here.