Both will address delegates gathered at the Time Warner Arena in Charlotte, N.C., on Sept. 4, two days before President Obama will formally accept the party's nomination at the nearby Bank of America Stadium.
Convention chairman Los Angeles Mayor Antonio Villaraigosa called the first lady an "inspiration to millions of Americans" who can offer a compelling perspective on her husband.
"The person who knows him best, she will offer unique insights into the President as a husband, father and a leader over the last four years," he said in a statement. She also spoke on the first night of the convention in 2008.
Villaraigosa said Castro, who will be the first Latino to deliver a keynote address at a Democratic convention, would speak to Obama's economic philosophy which he has embodied in Texas.
"As mayor, Julián Castro has worked tirelessly to move San Antonio forward by building its economy from the middle out, not the top down, by putting the city on a path to being a leader in the new energy economy and making innovative investments in education to prepare San Antonio's students for the jobs of the future," he said. "That's the same vision forward for the middle class the President has."
The announcement of Obama and Castro as speakers comes as Democrats seek to build pre-convention buzz. On Monday, organizers announced that Bill Clinton would deliver the formal nominating address on the second night of the convention. Former Obama adviser turned Massachusetts U.S. Senate candidate Elizabeth Warren will also speak.