During the 2000s, he appeared in both Mexican and American films including Frida, Open Range, Dirty Dancing: Havana Nights, The Terminal, Criminal, Milk, Sólo quiero caminar, and Rudo y Cursi.

In the 2010s, he co-starred in a variety of films like the sci-fi Elysium, comedy Casa de mi Padre, and the animated musical The Book of Life, before starring as Andor in 2016's Rogue One, a role he reprises in the 2022 Disney+ series of the same name.

Throughout his career, he has appeared in Mexican theater productions and produced a number of film and television projects, many with his close collaborator Gael García Bernal.

He is the creator and director of the 2013 Fusion TV docu-series Back Home, his Amazon Studios talk show Pan y Circo, which premiered in 2020, and the 2021 Netflix scripted series Everything Will Be Fine [es].

Luna starred as Tenoch Iturbide alongside Gael García Bernal in Y tu mamá también (2001), a Mexican road trip film directed by Alfonso Cuarón.

[13] In the following two years, Luna acted in Mexican films along with English-language productions like the miniseries Fidel, Vampires: Los Muertos, the Kevin Costner western Open Range, and the Academy Award-winning Frida.

He subsequently had starring roles in the films Fade to Black (2006) and Harmony Korine's Mister Lonely (2007), and also appeared in the critically acclaimed biopic Milk (2008).

In the early 2010s, Luna co-starred in the Will Ferrell Spanish-language comedy Casa de mi Padre (2012) and appeared in the action films Contraband and Elysium.

[18] Rogue One: A Star Wars Story premiered in 2016, and his portrayal of Cassian Andor,[19] a captain and intelligence officer of the Rebel Alliance, brought him international attention.

[23] In 2017, it was announced that he would star in a Scarface remake;[24] however, the film went through multiple rewrites and directorial changes in the following years, losing Luna as the lead in the process.

[25] Luna starred as Mexican drug cartel leader Miguel Ángel Félix Gallardo in the Netflix series Narcos: Mexico,[26] which premiered in November 2018.

With frequent collaborator Gael García Bernal and producer Pablo Cruz, Luna created the production company Canana Films in 2005.

[31] Luna and García Bernal departed Canana in 2018, subsequently founding a new production company, La Corriente del Golfo.

[29] In October 2019, it was announced that Luna and García Bernal were joining the Creative Advisory Board for the TV and film development company EXILE Content.

[43] It was announced in 2019 that Luna and García Bernal would executive produce an epic miniseries Cortes for Amazon starring Javier Bardem as conquistador Hernán Cortés, then described as "the largest Spanish-language production of all time.

In 2023, it was announced that Luna is set to executive produce and star in a 'The Boys: Mexico' spinoff alongside his frequent collaborator and friend, Gael García Bernal.