Maxi Lopez netted a 95th-minute winner to secure Neroverdi their first-ever win at the Juventus Stadium as the home side were exposed on the break.
Despite dominating the first period, Juventus couldn't find a way past the resolute visitors, who then found a breakthrough against the run of play through Davide Frattesi just before half-time.
Sassuolo had really upped the ante after the break and deservedly found the winner when Lopez caught them on the break to seal the winner in the final minute of stoppage-time.
Juventus were condemned to a third defeat of the season in just 10 games, the most they've suffered at this stage since the 2015-16 season.
Here are the player ratings for Juventus:
Mattia Perin - 6.5/10
He was beaten to Fratessi's opener and was kept busy by Sassuolo for the first 15 minutes or so of the second period, but Perin held his ground well.
Danilo - 6/10
His passing was first-rate but Danilo's forward runs didn't yield anything meaningful, failing to get a single cross away.
Leonardo Bonucci - 7/10
The veteran Juventus star showed tremendous calm and composure during Sassuolo's moments of pressure and looked to get the ball out of tight spaces too. His 400th Serie A appearance, however, was ruined by that late goal.
Matthijs de Ligt - 6.5/10
He couldn't prevent Sassuolo's opener but was otherwise assured and completed 96% of his passes too.
Mattia De Sciglio - N/A
His night, unfortunately, came to a premature end with an injury, trudging off the pitch as early as the 13th minute.
Federico Chiesa - 6/10
The playmaking wizard struggled to weave his magic tonight and was also unlucky to see a good effort ricochet off the bar.
Weston McKennie - 7/10
He got Juventus level with a perfectly-placed header but was largely anonymous in the game until then. Also tried his best to track Lopez on the counter but couldn't prevent the winner.
Manuel Locatelli - 6/10
A tough night for Locatelli against his old club, winning only three of his 12 ground duels and getting dribbled past five times, indicating a lack of pace too. Not at his creative best either.
Adrien Rabiot - 6.5/10
The Frenchman didn't always impress and was taken off at half-time.
Paulo Dybala - 7/10
Arguably the most dangerous Juventus player on the night, Dybala tested Sassuolo with a couple of shots, laid out three key passes, and also set up McKennie for the equalizer with a fine free-kick.
Alvaro Morata - 6/10
The Spaniard was a total non-factor today, struggling to get a single shot away and making only five accurate passes in 63 minutes of action!
Alex Sandro - 7/10
He replaced the injured De Sciglio in the opening stanza and put up a lively attacking performance. Defensively, though, he was poor.
Juan Cuadrado - 7.5/10
Cuadrado brought a lot of industry, creativity and menace to Juventus' right flank. He fought hard for the ball too, winning nine of his 12 ground duels, and laying three key passes.
Arthur - 5/10
The former Barcelona man was barely seen after coming on.
Dejan Kulusevski - 5/10
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He, too, vanished amid a sea of black and white shirts after stepping into the breach around the same time as the Brazilian.