The headquarters of the moderate Islamist Ennahda party, which rules in a fractious coalition with secularists, was set ablaze after Chokri Belaid, an outspoken critic of the government, was gunned down outside his home in the capital.
Prime Minister Hamadi Jebali, who said the identity of the attacker was not known, condemned Belaid's killing as a political assassination and a strike against the "Arab Spring" revolution. Ennahda denied any involvement by the part.
Despite calls for calm from the president, 8,000 protesters, massed outside the Interior Ministry, calling for the fall of the government, and thousands more demonstrated in cities including Mahdia, Sousse, Monastir and Sidi Bouzid, the cradle of the revolution, where police fired teargas and warning shots.
"This is a black day in the history of modern Tunisia ... Today we say to the Islamists, 'get out' ... enough is enough," said Souad, a 40-year-old teacher outside the Interior Ministry in Tunis. "Tunisia will sink in the blood if you stay in power."
The small North African state was the first Arab country to oust its leader and hold free elections as uprisings spread around the region, leading to the ousting of the rulers of Egypt, Yemen and Libya and to the civil war in Syria.
But like in Egypt, many who campaigned for freedom from repression under autocratic rulers and better prospects for their future now feel their revolutions have been hijacked by Islamists they accuse of clamping down on personal freedoms, with no sign of new jobs or improvements in infrastructure.
Since the uprising, the government has faced a string of protests over economic hardship and Tunisia's future path, with many complaining hardline Salafists were taking over the revolution in the former French colony dominated previously by a secular elite under the dictatorship of Zine al-Abidine Ben Ali.
Last year, Salafist groups prevented several concerts and plays from taking place in Tunisian cities, saying they violated Islamic principles, worrying the secular-minded among the 11 million Tunisians, who fear freedom of expression is in danger.
Declining trade with the crisis-hit euro zone has also left Tunisians struggling to achieve the better living standards many had hoped for following Ben Ali's departure. Any further signs of unrest could scare off tourists vital to an industry only just recovering from the revolution.
"More than 4,000 are protesting now, burning tyres and throwing stones at the police," Mehdi Horchani, a Sidi Bouzid resident, told Reuters. "There is great anger."
Jobless graduate Mohamed Bouazizi set himself on fire in December 2010 in the city, 300 km (180 miles) southwest of Tunis, after police confiscated his unlicensed fruit cart, triggering the "Jasmine Revolution" that forced Ben Ali to flee to Saudi Arabia less than a month later, on January 14, 2011.
President Moncef Marzouki, who last month warned the tension between secularists and Islamsits might lead to "civil war", cancelled a visit to Egypt scheduled for Thursday and cut short a trip to France, where he addressed the European Parliament.
"We will continue to fight the enemies of the revolution," the secularist leader told European Union lawmakers in Strasbourg.
Belaid, who died in hospital, was a leading member of the opposition Popular Front party. A lawyer and human rights activist, he had been a constant critic of the government, accusing it of being a puppet of the rulers in the small but wealthy Gulf state of Qatar, which Tunisia denies.
"Chokri Belaid was killed today by four bullets to the head and chest," Ziad Lakhader, a leader of the Popular Front, told Reuters. "Doctors told us that he has died. This is a sad day for Tunisia."
Ennahda Prime Minister Jebali said the killers wanted to "silence his voice".
"The murder of Belaid is a political assassination and the assassination of the Tunisian revolution," he said.
Party President Rached Ghannouchi denied any involvement in the killing. Belaid said earlier this week that dozens of people close to the government attacked a meeting of his party.
"Is it possible that the ruling party could carry out this assassination when it would disrupt investment and tourism?" Ghannouchi told Reuters.
He blamed those seeking to derail Tunisia's democratic transition after a 2011 uprising. "Tunisia today is in the biggest political stalemate since the revolution. We should be quiet and not fall into a spiral of violence. We need unity more than ever," Ghannouchi said.
He accused secular opponents of stirring up sentiment against his party following Belaid's death. "The result is burning and attacking the headquarters of our party in many areas," he said.
French President Francois Hollande condemned the shooting, saying he was concerned by the rise of violence in Paris's former dominion, where the government says al Qaeda-linked militants linked to those in neighbouring countries have been accumulating weapons with the aim of creating an Islamic state.
"This murder deprives Tunisia of one of its most courageous and free voices," Hollande's office said in a statement.
Riccardo Fabiani, Eurasia analyst on Tunisia, described it as a "major failure for Tunisian politics".
"The question is now what is Ennahda going to do and what are its allies going to do?" he said. "They could be forced to withdraw from the government which would lead to a major crisis in the transition."
Marzouki warned last month that the conflict between Islamists and secularists could lead to civil war and called for a national dialogue that included all political groupings.
Ennahda won 42 percent of seats in a parliamentary election in 2011 and formed a government in coalition with two secular parties, the Congress for the Republic, to which President Marzouki belongs, and Ettakatol.
Marzouki's party threatened on Sunday to withdraw from the government unless it dropped two Islamist ministers.