Google said it has filed a formal complaint against Microsoft with the European Union’s top antitrust regulator, escalating a long-running dispute over the cloud-computing business.

In the complaint, the search giant accuses Microsoft of abusing its market power in enterprise software to push businesses to use its Azure cloud platform—and keep them locked in there.

The antitrust complaint is a turnaround for Google, which nearly two decades ago became a sustained target of EU antitrust action in part stemming from complaints supported by Microsoft. The two companies for a time made peace , but that accord has unraveled in recent years . This is the latest broadside.

Microsoft said it had settled similar concerns with European cloud providers and expected that Google’s complaint would “fail to persuade the European Commission.”

Google, owned by Alphabet , has long trailed Microsoft and Amazon.com ’s Amazon Web Services in the cloud business. Google executives say its complaint is important now because many companies will be moving more of their work to the cloud in the coming years.

Any practical impact of Google’s complaint could take years to be felt. EU competition enforcers will have to decide whether to pursue the complaint, then whether to file charges and then decide on any potential infringements, fines and remedies—which themselves can be challenged in court. The process normally takes years.

For instance, it took the EU four years before filing charges in July against Microsoft over allegations stemming from a complaint workplace-messaging app Slack filed in 2020. A decision and any possible remedies in that case are pending.

Google’s complaint, which the company says it filed late Tuesday, comes after Microsoft in July settled a separate complaint from a trade group representing European cloud-services providers over similar issues. Under the deal, the trade group withdrew its complaint and Microsoft agreed to change some of its licensing practices while also reimbursing the group for its costs.

On Wednesday, Google criticized that deal, arguing it does little to lift what it says are barriers to customer choice, which it alleged has been restricted and worsening since Microsoft made changes in 2019 to its licensing practices. Google argues that customers using Windows Server that want to switch to the cloud are unfairly penalized if they want to use competing cloud services. Google says Microsoft’s licensing conditions limit businesses’ choice of cloud providers and raise their costs.

Google has also made similar arguments to the U.K.’s competition regulator, as part of that body’s market investigation into cloud services, saying that Microsoft’s restrictions keep it and others from competing for much of the enterprise cloud market.

“We believe this regulatory action is the only way to end Microsoft vendor lock-in…and create a level playing field for competitors,” said Amit Zavery , vice president and head of platform for Google Cloud.

Write to Sam Schechner at Sam.Schechner@wsj.com