The Kremlin said attacks on Luhansk, Donetsk, Zaporizhzhia and Kherson would in future be regarded as attacks on Russia, but refused to say whether it would retaliate with nuclear weapons.
Western leaders swiftly condemned the annexations as illegal. The US and Britain announced new sanctions, while Ukraine countered with a surprise announcement that it was applying to join Nato.
In a speech at an opulent Kremlin hall, Mr Putin said the four regions would not be up for discussion in any peace talks with Ukraine.
“People from Kherson, Donetsk, Luhansk and Zaporizhzhia become our citizens for ever,” he told dignitaries before he signed the acts of annexation.
“There is nothing stronger than the decisive will of millions of people who, according to their culture, to their language, believe they are part of Russia.”
Friday’s ceremony came after so-called referendums organised by Russian officials purportedly gave backing to the annexation in what Ukraine and its allies dismissed as a charade.
The four regions make up about 15 per cent of Ukraine’s internationally recognised territory. Russian troops occupy nearly all of Luhansk and Kherson, while Ukrainian forces have footholds in Donetsk and Zaporizhzhia.
In a combative speech filled with grievances with the West, Mr Putin:
· Said that Russia will defend its land “with all our strength and all our means”;
· Bemoaned the collapse of the former Soviet empire but said Russia was not seeking to restore the USSR;
· Defended the invasion of Ukraine despite Russian setbacks on the battlefield;
· During a diatribe against western hegemony, accused the US and its allies of being behind the mysterious Nord Stream gas leaks.
After signing the documents, Mr Putin shook hands with leaders of the four regions while the national anthem was played, before the five men clasped hands and chanted “Russia, Russia”.
“After the collapse of the USSR, the West decided that the world would forever have to put up with its diktats … but Russia has been reborn and strengthened,” Mr Putin said.
Nato and western powers said they would never recognise the land grab, described by Estonian Prime Minister Kaja Kallas as the most serious escalation of the conflict since Russia invaded Ukraine in February.
“Putin has mobilised hundreds of thousands of more troops, engaged in irresponsible nuclear sabre-rattling and now illegally annexed more Ukrainian territory,” Nato chief Jens Stoltenberg said in remarks on Friday.
“Together, this represents the most serious escalation since the start of the war. This land grab is illegal and illegitimate.
“Nato allies do not and will not recognise any of this territory as part of Russia. We call on all states to reject Russia's blatant attempts at territorial conquest. These lands are Ukraine's.”
US President Joe Biden described Mr Putin's actions as “phoney claims of annexation” and said Russia was “showing its contempt for peaceful nations everywhere”.
“Make no mistake: these actions have no legitimacy. The United States will always honour Ukraine’s internationally recognised borders,” he said.
Mr Biden and others promised they would remain undeterred by Russia's attempt to raise the stakes of a Ukrainian counter-attack.
“The strategy of seeking to falsely present Ukraine's territory as Russia's and purporting that the war may now be taking place on Russia's territory will not shake our resolve,” a joint statement from leaders of the EU said.
Vladimir Putin annexes four regions of Ukraine — in pictures
UN Secretary General Antonio Guterres called Russia’s move a breach of the UN Charter, while Britain’s Prime Minister Liz Truss said Mr Putin was showing “disregard for the lives of the Ukrainian people he claims to represent”.
“Putin cannot be allowed to alter international borders using brute force. We will ensure he loses this illegal war,” Ms Truss said.
Mr Biden said new sanctions would target those who provided political or economic support for Russia's attempted annexations. He said the US would continue to help Ukraine by “strengthening its hand militarily and diplomatically”.
Britain separately announced an export ban on hundreds of goods and said Russia would be denied access to services such as engineering, architecture and certain legal advice from the UK.
The measures are designed to bring the weight of London's powerful service sector to bear and “disrupt and degrade the capability of Russian businesses to keep pace in the international market”, the Foreign Office said.
Foreign Secretary James Cleverly said Russia “must be held to account for this abhorrent violation of international law”.
In Kyiv, Ukrainian President Volodymyr Zelenskyy mirrored Mr Putin's ceremonies by signing documents on Ukraine's Nato application. He said Ukraine was seeking an accelerated accession after proving itself up to Nato standards during the war.
“De facto, we have already made our way to Nato,” Mr Zelenskyy said after meeting defence and security chiefs on Friday. “Today, Ukraine is applying to make it de jure.”
Ukraine's long-standing ambition to join Nato is a prime grievance for Russia and was one of the reasons behind the invasion in February.
Donetsk and Luhansk together make up the coal-rich, largely Russian-speaking Donbas region where fighting has raged since 2014. Mr Putin cited the defence of Russian speakers there as one of his justifications for invading Ukraine.
Kherson lies just north of the Crimean peninsula annexed by Russia in similar fashion in 2014. Zaporizhzhia became the centre of international attention after shelling near a large nuclear power plant.
The move to annex the four regions comes a week after Mr Putin announced the partial mobilisation of Russian civilians, in what Ukraine’s allies described as a sign the invasion was going badly.
British intelligence reported on Friday that some of the newly mobilised reservists had been told to source their own medical supplies because of equipment shortages.
The European Union accused Russia of carrying out a “heinous attack” on a civilian convoy in southern Ukraine. Local authorities said 25 people were killed.