The 2016 Portland Timbers season was the sixth season for the Portland Timbers in Major League Soccer (MLS), the top-flight professional soccer league in the United States and Canada. The Timbers began this season as 2015 MLS Cup champions, but failed to qualify for the 2016 playoffs after a 4–1 away defeat to Vancouver Whitecaps FC on the final league day, also resulting in them failing to win the Cascadia Cup.

The reigning champions started their season against Columbus Crew SC. It was the fourth consecutive home opener being the first match of the season for the Timbers. As it has been since 2011, the Timbers Army led the national anthem and raised a tifo remembering the MLS Cup final victory with the words "Kiss. Dance. Love. You always remember your first." The Timbers were able to start strong with Valeri making the first goal in the 23rd minute from a free kick just outside the 18-yard box. Columbus would soon return the blow with a goal that came out of nothing. In the 68th minute, Higuaín was able to control the ball that lightly bounced off of Taylor's back and finish with a bicycle kick to equalize Columbus Crew SC. The Timbers answered in the 79th minute with a goal from Adi, who recovered the deflection from goalkeeper Steve Clark that blocked Asprilla's low shot. The Timbers won, 2–1.

The Timbers felt the loss of Liam Ridgewell who was ruled out with a hamstring injury shortly before the San Jose away match. Jermaine Taylor would be moved into the center back position due to Ridgewell's injury and give Zack Valentin his first start for the Timbers as he filled Taylor's position at left back. The Timber's would fall 2–1 with a late goal from substitute Jack McInerney.

Jack Barmby was announced to join the Timbers on loan from Leicester City and Darren Mattocks was signed from Vancouver Whitecaps FC.

The Timbers faced off with Real Salt Lake at home for a 2–2 draw. Despite playing against a 9-man Salt Lake squad, out-shooting them 26–7, and having 17 corners to their 4, the Timbers were unable to steal the victory and had to settle for the draw. Darren Mattocks was subbed in during this match, making it his first match since joining the Timbers. Fanendo Adi was the lone scorer, putting the ball in the back of the net in the 79th minute and converted a penalty in the 84th minute.

Position at the end of March

The Timbers started April on the road in Orlando, Florida, as they faced the current top team in the Eastern Conference, Orlando City FC. The team continued to tinker with the left back position, this time starting Jack Barmby instead of Valentin, giving Barmy his first start and appearance for the Timbers. The Timbers would suffer a harsh defeat of 4–1 with Fanendo Adi's penalty being blocked and allowing Orlando to convert their penalty. Jack McInerney was able to perform a late goal again, preventing Orlando's shutout.

The Timbers continued their search for a win on the road, this time against LA Galaxy. Chris Klute returned to the first team from Timbers 2, getting his first appearance as the squad continued to find a replacement for the left back position. The Timbers also changed their formation for this match from a 4–3–3 to a 4–2–3–1 putting Jack Jewsbury with Chara in the defensive midfielder positions. in the 52nd minute, Diego Valeri was able to send the ball to Fanendo Adi who managed a nice angle shot to send the ball inside the far end of the net. In the 74th minute, the Darlington Nagbe had to be carried off the pitch and return to the locker room via wheelchair due to a serious foul by Nigel de Jong to the ankle, who managed to escape with only a yellow card. Later on during the week, the MLS Disciplinary Committee would rule it was a red and de Jong was handed a 3-game suspension. Nagbe would later be ruled with an ankle sprain, contusion. Unfortunately, the Timbers would have to settle for the draw due to an own goal from Nat Borchers in the 83rd minute, continuing the Timbers search for a win on the road.