The shooting came after preliminary results showed Parti Quebecois (PQ) pipped the ruling Liberals in Tuesday's election and would have to be content with a minority government, effectively ruling out another referendum on breaking away from Canada.
Pauline Marois, newly elected as the first female premier of Quebec, had just told a rally the province would one day be independent when bodyguards rushed her from the stage. She later returned to finish her speech.
Such incidents are rare in Canada, where murder levels are around one-third of those in the United States and political violence is extremely rare.
Montreal police said a man entered the back of the Metropolis theatre just before midnight with a rifle and a handgun and shot two people. A man in his forties died immediately, while another was taken to hospital in a critical condition.
TV pictures showed police subduing a large man with a rifle who was dressed in a black cape and a black face mask. He appeared to shout in French the phrase "The English are waking up."
Ms Marois had promised to strengthen laws designed to ensure the dominance of the French language, which has worried some in the minority English-speaking community.
Carl Vallee, a spokesman for Canadian Prime Minister Stephen Harper, said: "We are appalled by this violence."
Security sources said Montreal police cordoned off a truck they suspected contained weapons. Other media outlets said the dead man was a technician at the theatre and the badly wounded man was a driver of the campaign bus.
The last political killing in Canada was in 1970, when a Quebec nationalist group kidnapped provincial Labour Minister Pierre Laporte and a British diplomat. Mr Laporte was found strangled.
Almost lost in the aftermath of the shooting was the fact PQ won 54 of the 125 seats, ending nine years of Liberal rule.
Previous PQ governments held independence referendums in 1980 and 1995, but both failed.
Ms Marois is promising another vote, but that could be years away. A recent poll showed only 28% of Quebecers back independence.
She said she would first concentrate on the economy, in particular tackling the province's debt, imposing higher tax and royalty rates on mining firms and making foreign takeovers of Quebec firms more difficult.
The results showed the Liberals won 50 seats, down 14.
The PQ won 31.9% of the vote, compared to 31.2% for the Liberals.
Francois Legault, leader of the third-placed Coalition for the Future of Quebec party, indicated he would back Ms Marois if she focused on fighting corruption and improving the financially strapped public healthcare system.
His party wants to freeze all talk of a referendum for a decade.