Jay Tabb came up with the winner for the West Londoners in the 58th minute, and they then hung on to record only their fourth away league win of the campaign.
Colchester right-back Pat Baldwin was the culprit with a poor back pass which substitute Scott Fitzgerald managed to intercept. Fitzgerald's goal-bound shot was blocked by keeper Aidan Davison but the rebound fell kindly for Tabb, who swept it into an empty net.
The Bees had dominated the first half, yet the goal actually came against the run of play.
The Essex hosts had struck the woodwork in the first half with a long-range shot from Gareth Williams, grazing the top of the bar, and they came even closer to a goal early in the second period.
Ironically it was Baldwin, later the villain, who was almost crowned a hero with a 20-yard shot that dipped over keeper Stuart Nelson, only to come back off the underside of the bar.
After Tabb's opening goal, Colchester rather lost their way, although they did mount a terrific late rally.
There was drama in injury time when their keeper Davison ventured up field for a corner. He was in the right place to meet Kevin Watson's delivery with a good header that had his opposite number Nelson beaten, only for his effort to be cleared off the goal-line.
It is Colchester's dreadful home form that is costing them dear this season. This is their ninth defeat from 18 league matches at Layer Road.
Their keeper Davison kept them in the match in the first half with a string of good saves as Brentford showed no signs of tiredness following their midweek defeat by Southampton in the FA Cup.
Impressive midfielder Ed Hutchinson was close to an early goal with a header that Davison did well to save, and the U's keeper again showed alertness when finger-tipping an inswinging corner by Stewart Talbot over the bar.
Colchester were happy to still be level at the break, and ironically it was when they were enjoying their best spell of the game that Brentford struck with the killer blow.