He took over from the regime's founding figurehead, Ayatollah Ruhollah Khomeini, when he died just over 10 years later, in June 1989.
Iran's supreme leader has the final say in all matters of state.
Therefore Khamenei's death, after almost 37 years in power, marks a major transition.
Iran latest: Ayatollah Khamenei killed
Which senior leaders have died?
In addition to Khamenei, several other senior officials were killed in US/Israeli airstrikes too.
They include Iran's army chief of staff, General Abdol Rahim Mousavi, and defence minister General Aziz Nasirzadeh.
Also killed was Major General Mohammad Pakpour, who took over as the Revolutionary Guard's top commander after Israel killed its last commander last June, and Ali Shamkhani, a top security adviser to Khamenei.
Iranian media said Khamenei's daughter, grandchild, son-in-law and daughter-in-law were killed as well.
The Israel Defence Forces also claimed it had killed Saleh Asadi, head of the Intelligence Directorate of the Khatam al Anbiya emergency command, Mohammad Shirazi, head of the military bureau, Hossein Jabal Amelian, head of SPND (Organisation of Defensive Innovation and Research) and Reza Mozaffari-Nia, a former head of SPND and former deputy defence minister.
What happens now?
A three-person temporary leadership council has been formed to govern the country, in line with Islamic Republic law.
It includes Iran's reformist president, Masoud Pezeshkian, and the hard-line head of the judiciary, Gholamhossein Mohseni Ejei.
There will also be a jurist, Alireza Arafi, who is a member of Iran's Guardian Council and head of the Basij, a volunteer paramilitary force.
Ali Larijani, Iran's head of security, said the council would be set up on Sunday.
"We had prepared for such moments and have plans in place for all scenarios, even for the time after the martyrdom of revered Imam Khamenei," said Mohammad Bagher Qalibaf, Iran's parliamentary speaker.
He added: "You'll see that after the leadership council is formed, the power and integrity of officials, defensive forces and the people will be beyond imagination."
Who chooses the new leader?
While the leadership council will govern in the interim, an 88-member panel called the Assembly of Experts will pick a new leader. Under Iranian law, that must happen as soon as possible.
The panel is made up of Shiite clerics elected every eight years and whose candidacies are approved by Iran's constitutional watchdog.
The Guardian Council is known for disqualifying candidates. It barred former Iranian president Hassan Rouhani from election to the Assembly of Experts in March 2024.
A relative moderate, he struck the JCPOA nuclear deal with world powers in 2015, from which the US, under Donald Trump, later withdrew.
Who could be the new supreme leader?
Under Iran's system of vilayat-e faqih - guardianship of the Islamic jurist - the supreme leader must be a senior leader with political and religious authority.
Khamenei's power was often wielded through close advisers. But it is unclear how many have survived, and he was never publicly recorded as naming a successor.
His son, Mojtaba Khamenei, a 56-year-old cleric, has been seen as a possible successor. He has never held government office, however.
That said, he has been described as a gatekeeper to his father.
He studied under religious conservatives in seminaries in the city of Qom, and is described as a hardliner with close ties to the Revolutionary Guard.
It had been thought that former president Ebrahim Raisi might seek the leadership, but he died in a helicopter crash in May 2024.
Regime change?
Donald Trump is urging Iranians to take the opportunity to overthrow the Islamic Republic, which has been accused of murdering tens of thousands of its own citizens in recent weeks.
The US president has described the death of Khamenei as the "single greatest chance for the Iranian people to take back their country".
And he has claimed that many people in the Revolutionary Guard, military and other security and police forces "no longer want to fight".
Read more:
Dubai hotels hit during Iranian missile fire
How have Iranians reacted to death of supreme leader?
Before the Iranian revolution Iran was ruled by a monarchy, with the king called the "shah".
Reza Pahlavi, the US-based son of the shah who was deposed in the 1979 revolution has said: "With (Khamenei's) death, the Islamic Republic has in effect reached its end and will very soon be consigned to the dustbin of history."
Any attempts to appoint a successor to Khamenei are "doomed to fail from the outset", Pahlavi added, claiming they will have neither longevity nor legitimacy.
He has urged Iran's military, law enforcement and security forces to take their "final opportunity to join the nation".
Iranian forces have launched hundreds of drones and missiles in multiple waves across the Middle East, but without inflicting significant harm against American assets in the region, such as sinking a warship or destroying a base.
By contrast, American and Israeli strikes have already devastated regime targets in Iran. They have taken out the head of the regime, Ali Khamenei, as well as the army's chief of staff, General Abdol Rahim Mousavi, and defence minister General Aziz Nasirzadeh.
Iran latest: Three US service members killed
Then again, it is only day two of the war and the Islamic Revolutionary Guard Corps (IRGC) has pledged "severe, decisive and regret-inducing punishment".
Yet the more time that passes without this rhetoric becoming a reality, the greater the questions about whether Iran's most feared military forces still have the capability to locate, target and strike the US and Israeli warships and jets attacking them.
General Sir Richard Barrons, a former senior UK military officer, said a number of factors would likely be limiting Iran's options for manoeuvre, not least the loss of so many top leaders.
Any move to fire missiles would also expose the launch site to American and Israeli attacks from the air, meaning Iran's missile launchers would have "quite a short" life expectancy.
In addition, previous attacks by both the US and Israel against Iran over the past couple of years have already degraded its missile stockpiles, launchers and air defences to blunt the regime's ability to detect incoming enemy aircraft.
All of this could explain why so few US and Israeli military targets appear so far to have suffered much meaningful damage despite Iran firing hundreds of missiles and drones.
Though the full extent of any damage is unclear.
UK Defence Secretary, John Healey, warned that a wounded Iran still has the capacity to cause harm - just potentially in even more erratic ways with little regard for the impact on the millions of civilians who live across the Gulf.
"This regime is lashing out. It's lashing out in an increasingly indiscriminate and widespread way," he said, speaking to Sky News's Sunday Morning with Trevor Philips.
"And people will be really concerned that it's not just military targets, but civilian airports like Kuwait, hotels in Dubai and Bahrain are being hit."
Tourist hotspots in the crossfire
Countries so far impacted include Israel, Qatar, the United Arab Emirates, Bahrain, Kuwait, Jordan and Iraq, where a number of American bases are located.
Many of the munitions appear to have been intercepted, but falling debris can be deadly.
With so much metal flying around, civilian and tourist locations have been caught in the crossfire, including one of the world's busiest airports in Dubai, where all flights have been halted, and the entrance of a luxury hotel.
This would have been terrifying for those affected but these strikes appear to have done nothing to degrade the ability of the US and Israel to keep hitting Iran.
Read more from Sky News:
How have Iranians reacted to death of supreme leader?
Trump has tipped Middle East into war that could last weeks
A high-value target for the regime must surely be two American aircraft carrier strike groups, led by the USS Gerald R Ford and the USS Abraham Lincoln.
They're thought to be located south of Cyprus.
An attempt to hit these warships might explain why the UK says two Iranian missiles were fired in the direction of the Mediterranean island. Britain has bases on Cyprus but they are not thought to have been the focus of the attack.
General Barrons said an outmatched Iran on the battlefield might seek alternative ways to strike back such as by closing the Strait of Hormuz - a vital transit point for global oil and gas exports.
Disruption to this shipping lane would impact economies around the world - and it is already starting with tankers being targeted, including off the coast of Oman.
President Trump is gambling that Iran lacks the capability to resist his overwhelming firepower. And that may well be the case.
But it only takes one Iranian missile penetrating American air defences to alter that calculation or at least dramatically increase the cost to Washington of its war.
Since the strikes on Saturday, Iran has carried out retaliatory attacks on Israel, US military installations around the Gulf, and Gulf Arab states, including the Saudi Arabian capital, Riyadh, and the global business and tourist hub of Dubai.
Politics latest: Healey refuses to rule out UK joining offensive action
Asked on Sunday Morning With Trevor Phillips if the UK's terror threat level is now under discussion, John Healey said: "Absolutely.
"When you get a regime like this lashing out in the Middle East indiscriminately and widely, hitting civilian as well as military targets, when you have some of its proxies capable of other actions on their behalf, then of course, our force protection in the region is at its highest.
"Our alert and our vigilance in the UK is also high."
Mr Healey said Home Secretary Shabanah Mahmood is responsible for making any announcement on the terror threat being raised in the UK.
"But, I've got full confidence in our intelligence agencies and our ability to deal with the terrorist threats," he added.
The current threat of a terror attack in the UK is "substantial", meaning an attack is likely.
On the terrorism threat rating system, substantial is in the middle, with "severe" and "critical" above it.
It has been at "substantial" since February 2022, when it was downgraded from "severe" (an attack is highly likely), where it was for four months following a suicide attack outside Liverpool Women's Hospital.
Threat levels are determined by the Joint Terrorism Analysis Centre, an independent body based within MI5.
Read more:
Iran latest: Israel carries out second day of strikes on Iran
Which Iranian officials were killed, who's now in charge and will be the new leader?
Mr Healey revealed to Trevor Phillips two missiles were fired from Iran towards Cyprus, where the UK has an RAF base.
He and Cyprus's president said they do not believe RAF Akrotiri on the island was the target.
There are currently two large American aircraft carrier strike groups off the Cyprus coast and it is believed they were the target.
The defence secretary also said the UK has taken down Iranian drones, and missiles landed on a base in Bahrain with 300 British personnel.
Allegedly hostile Iranian activities are suspected to have taken place on UK soil.
In 2023, a Chechen-born man was jailed after being found guilty of spying on a London-based dissident Iranian TV station to help terror plotters.
Magomed-Husejn Dovtaev, 31, was accused of conducting surveillance on the west London headquarters of Iran International as part of a plan by others to carry out a terror attack.
Dame Priti Patel, the Conservative shadow foreign secretary, told Trevor Phillips the government should "absolutely be concerned" about the security threat in the UK after the Iran strikes.
"The most important measure that they could take right now is the protection of our homeland. That is the duty and the responsibility of the government," she said.
"So they will be, I trust that they will be, absolutely looking at every single aspect of the threat levels and the security implications for our country."
Maritime activity has been almost brought to a standstill as the US and Iran have traded strikes.
Hundreds of tankers are usually travelling through the Strait of Hormuz between Persian Gulf and the Gulf of Oman at all times, with Iran to the north and Oman and the United Arab Emirates to the south. It's the only marine outlet for this region's main oil producers including Iran.
In 2024, around a fifth of all global oil was flowing through the narrow waterway - the equivalent of 20 million barrels a day.
A snapshot from a month ago on February 1 shows how busy the waterway is with vessels passing into and out of the waterway.
On February 28, the day after the US and Israel carried out their first strikes on Iran, far fewer vessels were in the area and very little movement. By March 1, very few ships were in the straight, and vessels appeared to cluster around large ports either side of the strait.
Sky's Data and Forensic team tracked several individual tankers. One - the KHK Empress - was already in the strait before turning back on Saturday at around 10:00 AM UTC. By Saturday evening, 4 others had turned away from the strait to head back out into the Gulf. And by Sunday they were all on the move out of the region.
Analytics agency Kpler estimates that these five ships have the capacity to carry around 10 million barrels of oil.
Fear of being targeted on the route are not unfounded. On March 1, a Palau-flagged oil tanker 'The Skylight' was attacked. Four people were injured and the whole crew of 20 people was evacuated.
The US navy is warning against navigation through the strait and some traders are suspending transit.
And interference to the ship tracking and communication system, AIS, is making the area even more dangerous. The images below show AIS signals, which ships use to broadcast their locations, on February 27 compared to 28 February.
The later image shows distorted signals, with ships broadcasting locations that appear to be far from their true positions, or even on land.
Volatility in the Gulf will have an impact across the world. Disruption here will in turn disrupt global markets and international trade.
The attacks, which are part of what the US has called Operation Epic Fury, killed Iran's supreme leader Ayatollah Ali Khamenei in his Tehran compound, as well as dozens of senior Iranian officials.
Read more:
Live updates on strikes
Which Iranian officials are dead?
Attacks close Middle East airports
The aerial assault triggered retaliatory strikes by Iran on multiple countries in the Middle East as the regime vowed revenge.
It has fired drones and missiles at Israel and US military installations around the Gulf, as well as the tourism and business hub of Dubai.
So what has happened and where?
Iran
Iranian authorities say more than 200 people have been killed since the start of the US and Israeli strikes on Saturday.
Donald Trump has told Fox News that 48 leaders in Iran have been killed, and said on his Truth Social platform that nine Iranian naval ships have been destroyed and sunk.
The American military said an Iranian Jamaran-class corvette was struck by US forces at Chabahar in the Gulf of Oman during the start of the operation.
Israel said it killed 40 top Iranian military officials, including defence minister Amir Nasirzadeh and Revolutionary Guards commander Mohammed Pakpour.
Iranian state media said an alleged strike on an all-girls school in the southern city of Minab on Saturday killed at least 165 people.
But the Israeli military said it was not aware of any Israeli or American strikes in the area of the school.
The Israeli military also said its planes have been carrying out strikes to open the "path to Tehran", and the majority of aerial defence systems in western and central Iran had been dismantled.
A spokesperson said many targets remained, including sites of military-industrial production.
Israel
At least nine people have been killed in Israel after a synagogue was hit by a strike in the central town of Beit Shemesh, according to authorities.
Another 28 people were wounded in the attack.
In Tel Aviv, loud explosions caused by missile strikes or interceptions could be heard.
United States
Three American service members have been killed and five have been seriously injured in military operations targeting Iran, the US military says. It did not identify the service members.
But Sky's US partner NBC News reported two US officials said the deadly attack happened in Kuwait.
United Arab Emirates
As Iran targeted the wider Gulf area, the United Arab Emirates (UAE) said on Sunday that three people had been killed so far in Iranian attacks on the country.
The defence ministry said Iran had launched 165 ballistic missiles targeting the nation, of which 152 were destroyed. Thirteen fell into the sea, it added.
Iran launched 541 bomb-carrying drones at the UAE, of which 506 were destroyed.
Shrapnel from Iranian attacks on the Emirates' capital of Abu Dhabi killed two people, state media said.
In Dubai, two people were injured after shrapnel from drones fell over two houses when they were intercepted, a Dubai state media office statement said.
Dubai's international airport, its landmark Burj Al Arab hotel and the Fairmont The Palm hotel on Palm Jumeirah Island all suffered damage, as did Abu Dhabi's international airport.
Kuwait
Kuwait's health ministry said on Sunday that one person has been killed and 20 people have been wounded in retaliatory attacks by Iran.
A dozen people were injured in Kuwait in previous attacks on Saturday.
The Kuwaiti defence ministry said Iran had fired 97 ballistic missiles and 283 drones towards the country.
Bahrain
Bahrain said a missile attack targeted the US Navy's 5th Fleet headquarters, and three buildings were damaged in the capital, Manama, and Muharraq city.
The UK's Defence Secretary John Healey said Iranian missile and drone strikes came within a few hundred metres of a group of 300 British military personnel in Bahrain.
Qatar
An RAF Typhoon jet operating from Qatar used an air-to-air missile to "successfully" shoot down an Iranian drone heading towards Qatari territory, a Ministry of Defence spokesperson said on Sunday.
Oman
A projectile hit the Marshall Islands-flagged product tanker MKD VYOM, killing a crew member on board as the vessel sailed off the coast of Oman, V.Ships - the company managing the vessel - has said.
Also in Oman, Iran struck the port of Duqm, which has been used by the US Navy as a logistical hub and is capable of hosting aircraft carriers.
Strait of Hormuz
The strategic Strait of Hormuz sees a fifth of the world's traded oil pass through it, making it a crucial trading route on Iran's southern border.
Oman says an oil tanker, a Palau-flagged vessel called Skylight, came under attack in the strait, wounding four mariners on board, the state-run Oman News Agency said.
A vessel off Mina Saqr in the UAE was hit by a projectile that caused a fire, according to the UK Maritime Trade Operations centre.
The blaze was extinguished and the vessel was set to continue on its way.
A vessel in the strait off Iran also came under attack after Iranian officials reportedly threatened vessels transiting the strait over the radio.
In a sign of disruption to energy supplies, at least 150 tankers including crude oil and liquefied natural gas vessels
dropped anchor in open Gulf waters beyond the Strait of Hormuz.
And dozens more were stationary on the other side of the chokepoint, shipping data showed.
Cyprus
UK Defence Secretary John Healey also said that two missiles were fired in the direction of Cyprus, where the UK has bases. "We don't believe they were targeted at Cyprus," he said.
Jordan
Jordan said it "dealt with" 49 drones and ballistic missiles.
Pakistan
At least nine people were killed in clashes between protesters and police after hundreds of people stormed the US consulate in the port city of Karachi.
Police and officials said at least 25 people were also wounded and some of them were in a critical condition. Police said the demonstrators were later dispersed and the situation was under control.