William Lewis said in a message to staff that it was the right time to leave and that "difficult decisions" had been made to ensure the paper's future.
The newspaper had announced on Wednesday that it was cutting a third of its workforce, dramatically scaling back its coverage of sport and international news.
The decision was condemned by many journalists and prompted criticism of the Post's billionaire owner, Jeff Bezos. Executive editor Matt Murray said the cuts would bring "stability".
Jeff D'Onofrio, who joined as the newspaper's chief financial officer last year, will serve as acting publisher and CEO, the Post said as it announced Lewis's departure.
A former Dow Jones chief executive and publisher of the Wall Street Journal, Lewis was appointed to the role at the Washington Post in 2023.
He has faced criticism from subscribers and employees as he tried to reverse financial losses at the daily.
Hundreds protested in front of the paper's headquarters in Washington DC on Thursday after the mass lay-offs, which included the paper's entire Middle East staff and its Kyiv-based Ukraine correspondent.
A GoFundMe page has been set up to support international employees hired locally or from subsidiaries outside the US - and therefore not eligible for protections afforded to members of the Washington Post Guild, the paper's union - who may be facing a sudden loss of housing, visas or benefits.
"Among those laid off are reporters in war zones living without electricity, breaking news hub reporters and editors in Seoul and London who cover atrocities around the world every day, correspondents who upended their lives to move overseas just a few months ago, and indispensable local staff without whom our journalism would be impossible," organiser Michelle Lee wrote on the page on Saturday.
"They are a dedicated, talented, multilingual, brilliant bunch. They don't deserve this."
The page had raised more than $180,000 (£130,000) out of a target of $200,000 as of Sunday evening.
A separate GoFundMe page organised by the guild for US staff has already raised more than $500,000.
Marty Baron, the Post's executive editor until 2021, said the cuts ranked "among the darkest days in the history of one of the world's greatest news organisations".
The departure of Lewis marks the latest upheaval for the leading US newspaper, which has seen a series of staff cuts and controversial editorial decisions in recent years.
Shortly before the 2024 US presidential election, Bezos, the founder of Amazon, broke with decades of tradition by deciding the newspaper would not endorse a presidential candidate.
The newspaper had endorsed a candidate in most presidential elections since the 1970s - all of whom had been Democrats.
The move caused widespread criticism and led to the loss of tens of thousands of subscribers.
Meanwhile, the opinion editor resigned in February last year when Bezos decided to focus the paper's comment section on "personal liberties and free markets".
Bezos, who acquired the newspaper in 2013, said pieces opposing those views would not be published.
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