The first half at White Hart Lane had seen little in the way of creativity from Spurs but they burst into life after an hour with new boy Lamela at the heart of their attacking prowess.
The 21-year-old has been used sparingly by Villas-Boas since his arrival from Roma and has yet to start a Barclays Premier League game for Spurs. However, he made the difference against the reigning Moldovan champions, who succumbed to a 2-1 defeat in north London.
Lamela put in one of his most assured performances in a Tottenham shirt and topped it off by tucking away the first goal of the game 15 minutes into the second half, before being tripped inside the box moments later to give Defoe a chance to make history from 12 yards.
The England striker converted the penalty to claim his 23rd European goal for the club, one more than Martin Chivers managed during his time at White Hart Lane.
But Spurs did not have everything their own way. For all of their defensive work, Sheriff threatened twice in the first half before Ismail Isa grabbed a goal on 72 minutes to keep Tottenham on their toes.
However, Spurs saw the game out for a 2-1 win to reach the knockout phase. Elsewhere, Michael Laudrup admitted he felt sorry for his players after seeing them again suffer the agony of conceding a stoppage-time equaliser as they drew 1-1 with ten-man Kuban Krasnodar in Russia last night.
The Swans were again undone in injury-time at Kuban Stadium as Ibrahima Balde cancelled out Wilfried Bony’s opener for the hosts, who had defender Xandao sent off with 15 minutes remaining.
The result means Swansea have now won just twice in their past nine games in all competitions. Laudrup was pleased with the response from his players after Sunday’s derby defeat to Cardiff, but just felt they had lacked a bit of good fortune.
“We were very pleased to see the reaction and the performance in the first half, we played some good football and there were good individual performances,” said the Dane. “We had good chances and went in front and it was quite an open game. I thought we did well.
“In the second half we tried to play a little more on the counter and had chances but the goal came.
“It was a ball we had to clear, but it was a good pass and a great finish. In the end we learned from the first game and we kept the ball so we cannot say anything about that.
“I feel sorry for the players because I think they deserved to win this game. But sometimes you need a little luck.”
Wigan lost 1-0 at Rubin Kazan in Group D but may have taken at least a point. The only goal of the game at the Central Stadium came in the 22nd minute when Oleg Kuzmin latched on to a lofted ball into the area and cracked it home.
Valencia and Fiorentina left it late to seal their places in the knockout stages with two games to spare.
The Spaniards came from behind to beat ten-man St Gallen 3-2 with two late goals in Switzerland, while Fiorentina scored twice in the last four minutes to secure a 2-1 win over Pandurii Targu Jiu in Romania.