WASHINGTON — The acting director of the F.B.I. contradicted the White House on two major issues on Thursday: the support of rank-and-file agents for the fired F.B.I. chief James B. Comey and the importance of the agency’s investigation into Russian election interference.

In a striking repudiation of official White House statements, the acting director, Andrew G. McCabe, said the inquiry was “highly significant” and pledged to the Senate Intelligence Committee that the F.B.I. would resist any attempt to influence or hobble the investigation.

“Simply put,” he said, “you cannot stop the men and women of the F.B.I. from doing the right thing.”

That Mr. McCabe felt compelled to assert the F.B.I.’s independence was itself remarkable, a byproduct of the unusually public effort by Mr. Trump and his aides to take focus off the investigations into Russia’s election meddling. He also said the F.B.I. investigation had the resources it needed, partly disputing an account that Mr. Comey had sought more aid.