Comandante Ramona, a diminutive Tzotzil Maya woman and the first Zapatista rebel to appear publicly in Mexico City after a brief but bloody uprising in 1994, died on the way to hospital in the southern state of Chiapas, said Marcos, the Zapatista chief, on Friday.
"Mexico has lost one of those fighters that matter, and a piece of our hearts has been ripped out," Marcos told supporters during a speech in Chiapas.
Marcos left his jungle hideout last Sunday to start a nationwide tour that seeks more support for Indians and the poor before July's presidential election.
He put the tour on hold to attend her funeral in the rebel stronghold of Oventic.
Women's rights champion
Always seen in the black ski mask worn by Zapatista leaders to protect their identity, Ramona was one of the main promoters of women's rights in the rebel group, which has ditched armed struggle in favour of political activism.
The rebels burst out of the jungle on New Year's Day 1994, taking over towns and attacking army positions in Chiapas,
Ramona (R), seen with Zapatista
chief Marcos in 1996
Mexico's poorest state, in clashes that claimed about 150
Ramona won the hearts of many Mexicans after media showed fellow Zapatista commanders towering above her during a first round of peace talks with the government in February 1994.
There has been no fighting in recent years but the Zapatistas control scores of small villages across Chiapas, which they govern under a separate system of laws based on Indian traditions.
Many senior Zapatista commanders are women.
Ramona received a kidney from her brother in a 1996 transplant paid for by supporters who sent in cash after an internet campaign.
Along with Marcos figures, dolls of Ramona complete with rifle and ski mask are popular with tourists visiting Chiapas.