Dreyfuss was born in Brooklyn, New York, the son of Geraldine Dreyfus (née Robbins; 1921–2000), a peace Activist, and Norman Dreyfus (1920–2013), an attorney and restaurateur, and was raised in the Bayside area of Queens, New York. His family is Jewish. He has commented that he "grew up thinking that Alfred Dreyfus and [he] are from the same family." His Father disliked New York, and moved the family first to Europe, and later to Los Angeles, California, when Dreyfuss was nine. Dreyfuss attended Beverly Hills High School.

Dreyfuss began acting in his youth, at Temple Emanuel of Beverly Hills Arts Center and Westside Jewish Community Center, under drama Teacher Bill Miller. He debuted in the TV production In Mama's House, when he was fifteen. He attended San Fernando Valley State College, now California State University, Northridge, for a year, and was a conscientious objector during the Vietnam War, working in alternate Service for two years, as a clerk in a Los Angeles hospital. During this time, he acted in a few small TV roles on shows such as Peyton Place, Gidget, That Girl, Gunsmoke, Bewitched, The Ghost & Mrs. Muir, and The Big Valley. During the late 1960s and early 1970s, he also performed on stage on Broadway, Off-Broadway, repertory, and improvisational theater.

Dreyfuss's first film role was a small, uncredited appearance in The Graduate. He had one line, "Shall I get the cops? I'll get the cops". He was also briefly seen as a stage hand in Valley of the Dolls (1967), in which he had a few lines. In 1973 he starred in the CBS pilot Catch-22. He appeared in the subsequent Dillinger, and landed a role in the 1973 hit American Graffiti, acting with other Future stars such as Harrison Ford and Ron Howard. Dreyfuss played his first lead role in the Canadian film The Apprenticeship of Duddy Kravitz (1974), receiving positive reviews, including praise from Pauline Kael.

Dreyfuss appeared in the play The Time of Your Life, which was revived on March 17, 1972 at the Huntington Hartford Theater in Los Angeles, and directed by Edwin Sherin.

Dreyfuss went on to star in the box office blockbusters Jaws (1975) and Close Encounters of the Third Kind (1977), both directed by Steven Spielberg. He won the 1978 Academy Award for Best Actor at the 50th Academy Awards ceremony for his portrayal of a struggling actor in The Goodbye Girl (1977), becoming the youngest actor to do so (at the age of 30 years, 125 days old), besting Marlon Brando, who had won his first Oscar in 1955 at the age of 30 years 360 days old. This record stood for 25 years until it was broken in 2003 by Adrien Brody, who was three weeks shy of age 30 at the time of the 75th Academy Awards ceremony.

Dreyfuss won the Academy Award for Best Actor in 1977 for The Goodbye Girl, and was nominated in 1995 for Mr. Holland's Opus. He has also won a Golden Globe Award, a BAFTA Award, and was nominated in 2002 for Screen Actors Guild Awards in the Outstanding Performance by a Male Actor in a Drama Series and Outstanding Performance by a Male Actor in a Television Movie or Miniseries categories.

Around 1978, Dreyfuss began using cocaine frequently; his addiction came to a head four years later in 1982, when he was arrested for possession of the drug after he blacked out while driving, and his Mercedes-Benz 450 SL struck a tree. He entered rehabilitation and eventually made a Hollywood comeback with the films Down And Out In Beverly Hills in 1986 and Stakeout the following year. Dreyfuss also starred in the Rob Reiner movie Stand by Me, a 1986 coming-of-age drama/comedy film adapted from Stephen King's novella The Body. Dreyfuss plays the elder Gordie Lachance, who narrates the film. In 1988, he reunited with Director Paul Mazursky to star in the political farce Moon Over Parador.

Dreyfuss married Writer and Producer Jeramie Rain in the early 1980s. With her, he had three children: Emily (born 1983), Benjamin (born 1986), and Harry (born 1990). His elder son, Benjamin, was born with Peters Anomaly, a rare genetic eye disorder which, after many operations, left him blind in his left eye. Dreyfuss and Rain have continued to raise money for ophthalmology centers throughout the United States. After his 1995 divorce from Rain, Dreyfuss married Janelle Lacey in 1999, but they divorced in 2005.

In 1989, Dreyfuss reunited with Spielberg on Always, a remake of A Guy Named Joe in which he co-starred with Holly Hunter. He had a starring role opposite Bill Murray in the 1991 comedy What About Bob?, as a Psychiatrist who goes crazy while trying to cope with a particularly obsessive new patient. That same year, Dreyfuss produced and starred as Georges Picquart in Prisoner of Honor, an HBO movie about the historical Dreyfus Affair.

In 1994, he participated in the historic Papal Concert to Commemorate the Shoah at the Vatican in the presence of Pope John Paul II, Rav Elio Toaff, chief rabbi of Rome, and Oscar Luigi Scalfaro, President of the Italian Republic. He recited Kaddish as part of a performance of Leonard Bernstein's Third Symphony with the Royal Philharmonic Orchestra under the baton of Gilbert Levine. The event was broadcast worldwide.

In 1995, Dreyfuss co-authored with science-fiction Writer Harry Turtledove the novel The Two Georges, a steampunk/alternate history/mystery piece set in the year 1995 of a timeline where the American Revolution was peacefully avoided. Thomas Gainsborough's painting of George Washington and King George III, which symbolizes English-speaking North Americans' loyalty to the British Empire, is stolen by anti-Imperial terrorists, and officers of the Royal American Mounted Police must find it before it is destroyed.

Dreyfuss recorded the voiceover for the Apple Computer "Think Different" ad campaign in 1997.

Dreyfuss has been outspoken on the issue of how the media influences the shaping of public opinion, policy, and legislation. In the 2000s, he expressed his sentiments in favor of right to privacy, freedom of speech, democracy, and individual accountability.

In November 2004, he was scheduled to appear in The Producers in London, but withdrew from the production a week before opening night. The media noted that Dreyfuss was still suffering from problems relating to an operation for a herniated disc in January, and that the part of Max Bialystock in the play is a physically demanding one. Both he and his assistant for the production stated that Dreyfuss was accumulating injuries that required him to wear physical therapy supports during rehearsals. Dreyfuss was eventually fired from the production, though he would return in 2008.

Dreyfuss and Russian-born Svetlana Erokhin married in 2006 and lived in San Diego, California, although they frequently visited New York City and London, where Dreyfuss once lived. They also lived in Carlsbad, California. In February 2008, they bought a $1.5 million house in Encinitas, California, and plan to renovate the 1970s structure with state-of-the-art green technologies.

In early 2009, he appeared in the play Complicit (directed by Kevin Spacey) in London's Old Vic theatre. His participation in the play was subject to much controversy, owing to his use of an earpiece on stage, reportedly because of his inability to learn his lines in time. He guest-voiced as himself in the "Three Kings" episode of Family Guy in 2009, and later appeared again in the episode "Peter-assment". Dreyfuss has guest starred in the sixth season of Weeds as Warren Schiff, Nancy's high school Teacher to whom she had lost her virginity.

On June 10, 2011, Dreyfuss was made a Master Mason "at sight" by the Grand Master of Masons of the District of Columbia at the Washington DC Scottish Rite building, as well as a 32nd Degree Scottish Rite Mason. That evening he spoke at a banquet celebrating the 200th Anniversary of the Grand Lodge of the District of Columbia, about the Dreyfuss Initiative, a Research Society promoting civics and enlightenment values to be headquartered in Charleston, West Virginia.

Dreyfuss was among 99 other stars at the 2012 Academy Awards - Night of 100 Stars. He did an interview for the Bill Zucker Show with actor/singer Bill Zucker.

In 2014 he appeared with best-selling Lincoln scholar Ronald C. White in a documentary entitled "Lincoln's Greatest Speech", highlighting Lincoln's Second Inaugural Address, appearing as host of the program and reciting Lincoln's speech on camera.

On February 18, 2015, it was announced that Dreyfuss would portray Bernie Madoff in an upcoming miniseries. The first episode was telecast on February 3, 2016, co-starring Blythe Danner.

In 2017, Writer Jessica Teich accused Dreyfuss of exposing himself to her and attempting to get her to perform fellatio on him during the filming of the 1980s ABC comedy special Funny, You Don't Look 200: A Constitutional Vaudeville; in an interview with vulture.com, she said "I remember my face being brought close to his penis. I can’t remember how my face got close to his penis, but I do remember that the idea was that I was going to give him a blow job. I didn’t, and I left". Dreyfuss responded with a written statement denying that he had ever exposed himself to Teich. He did, however, state that he had been overly flirtatious in his past, and that he regretted that behavior, but he emphasized that he "value[s] and respect[s] women" and is "not an assaulter."