The chant has been heard at Chelsea's recent matches against Nottingham Forest and Manchester City, and also at the Manchester United v Everton FA Cup match, where it was aimed at the Toffees boss Frank Lampard, a former Chelsea player and manager.
While the FA has always condemned the use of the term, it has never felt able to charge clubs over its use in the past.
However, the PA news agency understands the recent conviction of Liverpool fan Paul Boardman, who admitted using the term on his way to last season's FA Cup final against Chelsea at Wembley, was a game changer, and opens the door for the FA to sanction clubs where fans are proven to have used the term.
An FA statement said: "Today, the FA has formally written to all clubs across the Premier League, EFL, National League, Women's Super League, Women's Championship and Steps 2-4, to remind them that it can pursue formal disciplinary action against any club whose supporters engage in discriminatory behaviour, now including the use of the term 'rent boy'.
"This important step follows the recent successful prosecution of an individual by the Crown Prosecution Service for homophobic abuse, specifically relating to the term 'rent boy'.
"The FA has now informed all clubs that it considers the 'rent boy' chant to be a breach of the FA rules. These rules apply to the conduct of supporters at both home and away fixtures, and clubs at all levels of English football have a responsibility to ensure their spectators behave appropriately when attending matches."
The letter is understood to be intended as a 'line in the sand' to clubs, so that any 'rent boy' chants from this point forwards could result in disciplinary action.
The Crown Prosecution Service said last week it was looking into reported use of the term by Forest fans during their New Year's Day match against Chelsea at the City Ground.
Chelsea Pride, an LGBTQ+ supporters' group, tweeted on the day of the game, calling on broadcaster Sky Sports to call out the chanting on air, adding: "This is now classed as a hate crime."
Forest's own LGBTQ+ Trickies group said it was "embarrassed and ashamed" by the abuse and apologised on behalf of those responsible.
There were 106 reported incidents of hate crime involving sexual orientation at matches in England and Wales during the 2021-22 season, according to Home Office figures released last year. That represented a 186 per cent increase on 2018-19, the last full season unaffected by the Covid-19 pandemic, when there were 37 such incidents reported.