General John Allen, commander of the International Security Assistance Force (Isaf), relieved Fuller as deputy commander of the effort to train Afghan security forces after Fuller told Politico that Afghan leaders were "isolated from reality", a US defence official said.
Pentagon spokesman George Little had said on Friday that defence secretary Leon Panetta was aware of the remarks and Fuller had been speaking for himself, not the US defence department.
"The secretary has full trust and confidence in General Allen's judgment with respect to his decision in this case," Little said in response to Allen's decision to relieve Fuller of his duties.
Speaking in a Politico interview that ran on Thursday, Fuller depicted Afghan officials as detached and unappreciative of American sacrifices and financial contributions to Afghanistan after 10 years of war.
The interview painted Fuller as critical of Karzai's recent comments suggesting Afghanistan would side with Pakistan if it went to war with the United States.
"Why don't you just poke me in the eye with a needle! You've got to be kidding me - I'm sorry, we just gave you $11.6bn and now you're telling me, 'I don't really care?'"
The interview quoted Fuller as saying Afghanistan did not recognise the sacrifice in "treasure and blood" the US was making for its security.
In July 2010 Barack Obama fired General Stanley McChrystal, then commander of US and Nato forces in Afghanistan, over remarks he and his aides made in an explosive Rolling Stone magazine article that disparaged the president and other civilian leaders.
While Fuller's job was far less senior than McChrystal's, the training of Afghan security forces has become more and more central to Nato's mission in Afghanistan as foreign forces gradually seek to put Afghan soldiers and police in charge of security.
Afghan security forces are far more numerous than they were and better skilled, but they still have inadequate fighting skills, poor equipment and widespread illiteracy.
While Obama plans to remove the 33,000 extra troops he sent following a 2009 review of Afghan war strategy, security conditions remain troubling. The United Nations says violence is at its worst level since the war began in 2001.
