Kightlinger was born and raised in Jamestown, New York. After touring as a stand-up comic, she got her break writing for the television sitcom Roseanne. She has written for Dennis Miller Live and for the 20th season (1994—1995) of Saturday Night Live. During that season, she impersonated several celebrities, including Brooke Shields, Marcia Clark, Paula Poundstone, Mary Hart, and Connie Chung.

Kightlinger worked as a writer and consulting producer on Will & Grace, and had a recurring guest role on the series as Sheila, a nurse at a fertility clinic. She was a correspondent on The Daily Show in 1998. She was a series regular in the HBO sitcom Lucky Louie. She created, wrote and starred in her own show on IFC, The Minor Accomplishments of Jackie Woodman, which brought her a Satellite "Best Actress" nomination.Kightlinger's book, Quick Shots of False Hope, which is described in The New York Times Book Review as "memorable, disturbing and darkly comic," is currently being adapted for film.As a filmmaker, Laura's short 2001 film, "Dependable People" won the Black Maria Director's Citation and the IFCT Best Director Award in 2002. Her documentary, "60 Spins Around the Sun" about a political satirist turned activist won 'Best Documentary" at the Empire State, Beverly Hills and Boston Film Festivals, and is available on Netflix. Her horror mockumentary, Cat Demon: Re-Exhumed is currently making the film festival rounds, having premiered in the LA Film Fest.

Rejection is my petrol. It’s what keeps me going. Laura Kightlinger's Major Accomplishment - Cable Shows - Page 2 | CRAVEONLINE.COM

In 1999, Kightlinger's book, Quick Shots of False Hope, a collection of short autobiographical stories was released.

In 2001, Kightlinger appeared in Shallow Hal alongside Gwyneth Paltrow.

She created, wrote, executive produced and starred in The Minor Accomplishments of Jackie Woodman, a comedy series about two best friends maneuvering through the not-so-glamorous side of Hollywood. The series aired on IFC from August 4, 2006 through September 23, 2007.

In 2003, Kightlinger directed a documentary about the Rockefeller Drug Laws, 60 Spins Around the Sun, which deals with Randy Credico, political satirist turned activist, and his fight to repeal the Rockefeller drug laws. The film follows Credico to Tulia, Texas during a racially motivated drug bust, which is the subject of an upcoming film starring Halle Berry. Included in this documentary are stand-up comics Larry David, Colin Quinn, Don Gavin, Vanessa Hollingshead and Nick DiPaolo.

"60 Spins Around the Sun" won 'Best Documentary' at the (2006) Empire State, Beverly Hills and Boston Film Festivals and is available on Netflix.com.

Kightlinger is a 1982 graduate of Southwestern High School in Jamestown, NY and a 1986 Emerson College graduate.