The leader of the wealthy northeastern region signed a decree on Saturday calling the vote on Nov. 9 and putting his local government on a collision course with Madrid.
"Neither the object nor the proceedings of the vote are compatible with the Spanish constitution," Prime Minister Mariano Rajoy said as he announced the legal challenge on television.
While it could take years for the constitutional court to rule on the issue, the vote will be effectively suspended as soon as the court decides to accept the case.
The court said it would hold an emergency meeting on Monday, with proceedings expected to start immediately.
Catalonia, which accounts for around a fifth of Spain's economy, has its own language and distinct culture and has long fought for greater autonomy, Reuters reported.