In the Ace Ventura movie, Spike appears in exactly one scene, where he apes Ace's movements. In the cartoon, he became Ace's full-on sidekick.
Cousin Itt is a frequently recurring character in the 1992 Addams Family cartoon when he only appeared on a few occasions in the original live-action show.
The Earl of Lemongrab is becoming this for Adventure Time. He got one appearance in season 3, one in season 4, three in season 5, and already has one in season 5.2 (keep in mind that this was the fifth episode of season 5.2).
In Avatar: The Last Airbender, Suki started out as an extremely minor character, appearing in only one episode of season 1, but as the series went on, her prominence gradually increased, culminating in her addition to the main cast in season 3.
Jinora in The Legend of Korra is a minor character in season 1, only really being one of Aang's grandkids. She serves vitally in the season 2 finale and by the end of season 3 is an Airbending master.
Jules and Verne, Doc Brown's kids in Back to the Future: the animated series, likely qualify for this, since their only appearance in the original films is a brief, non-speaking appearance at the end of the third film.
Harley Quinn was originally just a one-time moll character who made such an impression she became the Joker's pseudo-girlfriend/top henchwoman, then began developing relationships with other characters and got her own spotlight episodes, including a comic tie-in detailing her origins that got adapted into an episode. Then she became a Canon Immigrant into the comic universe and a Breakout Character in general. So she's ascended twice from one-shot to supporting character, from cartoon to comics, in that order.
Robin was featured infrequently in the first season, with the series bible even stating that he was not intended to be Batman's full-time partner like in the comics. Thanks to some Executive Meddling, the show was renamed The Adventures of Batman and Robin in its second season, and Robin appeared in nearly every episode.
Batgirl also appeared relatively infrequently throughout the original series, but was promoted to lead character status when the show was retooled as The New Batman Adventures. Bruce Timm claims her increased prominence was requested by network executives, who felt adding a woman to the main cast would win over female viewers.
A stated goal of Beware the Batman is to utilize obscure Batman villains who haven't been as heavily exploited as his A-list rogues. This means unknown (to the general public) villains like Magpie, Anarky, and Professor Pyg are getting their moment in the limelight, with Ra's al Ghul as the only remotely-recognizable villain confirmed for now. Katana also gets promoted to Batman's principal sidekick.
In the original 1993 Biker Mice from Mars series, Throttle's girlfriend General Carbine and the experienced Martian Freedom Fighter known as Stoker were minor characters who only appeared in a few episodes. In the 2006 revival, however, they are more prominent characters, with the Myth Arc having the Biker Mice answer to Carbine on their mission to retrieve the regenerator, a device Stoker invented that could be used to rehydrate Mars and save it from the drought it suffers.
Valerie Grey originally appeared briefly in the second episode of Danny Phantom, but after her popular life was ruined by ghosts (or rather, unintentionally by Danny), she became one of the most badass humans in the series, as well as one of Danny's toughest antagonists (though, in light of the Series Finale, she may now be an ally).
Daria started as a minor background character on Beavis And Butthead. She's mostly just seen in the classroom and we never meet her family (though her surname Morgendorffer is established and surprisingly remembered by the writers, even if the spelling isn't). The episodes where she speaks can be counted on one hand and even then, no episode revolved around her. Her interaction with the boys was mostly just her snarking at their stupidity and Beavis using his nickname for her, "Diarrhea," before she writes them off as morons and walks off.
Scruffy the Janitor. He shows up to comment on things when all other characters are used up. He's also occasionally featured as being even lazier than Fry, spending all his time in the basement reading porno magazines.
A strange case of this occurred with the "Number 9 Guy", who appears in many crowd shots as early as the series pilot, before finally getting a plot relevant role. The writers had always wanted to feature him, but were unable to fit him into an episode plot until "Into the Wild Green Yonder." Originally, he was to have been a part of a futuristic caste system, but this was abandoned early on.
The sexy young doctor named Dr. Cahill, (although Fry just called her Dr. Good and Sexy) who, after the first movie, become a regular character in the next seasons.
Harvey Birdman, Attorney at Law gives expanded roles to several Birdman villains, including Mentok, Reducto, Birdgirl (who started off brainwashed) and X. All had originally appeared in only one episode each.
Angel (a.k.a. Experiment 624) from Lilo & Stitch: The Series, to the point where in Japan she is featured with Stitch in most merchandise, overshadowing Lilo.
Charles Foster Ofdensen of Metalocalypse was just the routinely ignored voice of reason, and general straight man during early episodes. Then came the season 1 finale, where we learn he's a secret asskicker. Since then, he's become one of, it not, the most popular character on the show.
In Moral Orel, there's a fat woman who's often seen at church as well as in the background in many other scenes (including being picketed by Miss Censordoll's group of protesters). One of the last episodes, "Sundays" is all about her (spread out chronologically over the course of the show), revealing that her name is Florence, she's Officer Papermouth's ex-wife and that she has a crush on Reverend Putty.
Derpy Hooves is an excellent example of this. She was so unimportant that she almost didn't exist all together. She was present, with a normal appearance, among dozens of other background ponies in a longer sequence in the opening episode, but in one brief "blink and you'll miss it" cutaway shot, she was shown wall-eyed with a goofy grin. Word of God admits her momentary appearance was either an animation error or a joke by one of the artists. Despite that, the fans loved her. No, that's an understatement. They adored her. Story after story and cameo after cameo were created for her. Fans fleshed out her job, her personality, and even how she talked. Eventually, the show's creators caught on. Derpy slowly gained more acknowledgement throughout Season 1 and the first half of Season 2, first by the animators going back and derping her eyes in every scene she was in, then by placing her somewhere in the background of every new episode, then by giving her her own Funny Background Events, until finally it climaxed in the fourteenth episode of Season 2, where she was referred to by her Fan Nickname, given a voice actor, and interacted with a main character. Not bad for a animation error.
Of course, this led to a massive case of Double Standard when people wrote in, complaining that a girl should never be like how Derpy was and that she was supposedly highly offensive to the mentally disabled. Her name was cut out of the dialogue because it was assumed by the complainers to be some sort of slur, her wall-eyes were "fixed" in some shots, and her voice was changed note in the original version of the episode, her voice actor had gotten confused and thought she was a colt, so that last one was a legitimate correction. Granted, there was a Word of God moment where one of the writers expressed regret over doing it like that because of how her own child was. Didn't placate the fans and, eventually, she had a rather triumphant return along with a promise from Hasbro's vice-president that she here to stay.
Trixie. She began as a one-shot Villain of the Week, but received a massive amount of fan attention. Since then, she got a return episode in Season 3, another in Season 6, and is the focus of two story arcs in the comics as well as cameoing repeatedly. Her human counterpart in the Equestria Girls films is also a recurring supporting character.
Maud Pie. Of Pinkie's sisters, she alone gets to be the main focus for an episode and get multiple apparitions (also a cameo scene in Equestria Girls).
Cross-generation, there's Applejack. G1 Applejack: was the one who happened to be on the rickety bridge when it broke so the Sea Ponies could be introduced. Appeared once. Was more important in the comics, with the clumsiness and greed for apples that gave us the "Who's A Silly Pony" song and therefore meme, but except for one A Day in the Limelight story, had nothing to her but being a pony who really, really liked apples, at least in the issues currently available online. G3 Applejack: Background pony. Being colored lighter than her better-known toy color scheme means you'll have a hard time spotting her few blink-and-miss moments. FIM AJ: One of the main characters, part of a huge extended family, and with several episodes devoted to her. Her little sister is a secondary character, which means in an episode focused on the kids, if one adult is going to get a major role, there's a 50-50 chance it's her.
Twilight, too, though to a lesser degree than AJ. In the G1 cartoon, she only appeared once, and only had a few moments, but those moments were memorable and one was very important. In the comics, this is magnified: her appearances are not the most frequent, but when she does appear, you remember it.note In this case, she's insanely overpowered, and only appears at night, most often hiding herself in a magical fog when she must travel during the day. She's one of the most memorable things about the comic. But in terms of screentime, there are several downright "Who's that, again? Are you sure she was in the comics?" characters who actually got more. There was another Twilight, a pegasus who also appeared once.note How much of Twilight Sparkle is owed to her is unknown. Pegasus Twilight is purple, but so is the G3 earth pony Twilight Twinkle, who really looks like the inspiration for Twilight Sparkle in terms of design. Pegasus Twilight's cutie mark is a candle; an identical one adorns the door of Twilight Sparkle's library. Twilight Sparkle is, of course, the main character of the FIM series, and eventually goes on to become the fourth alicornprincess.
Stacy Hirano. In the early episodes, she appeared only infrequently, if at all (she was originally just an unheard voice on the other end of Candace's cell phone conversations). As the series progressed, she appeared much more often, and has actually played a role in the plot of some episodes ("Put that Putter Away", "Elementary my Dear Stacy", "Phineas & Ferb-busters", "The Lemonade Stand"). In fact, it was almost rare to see a current episode where she doesn't appear, even if it's only for a scene or two.
Norm. Started out as one of Doof's inventions (after learning "the enemy of the platypus is man", he built a robot man), and then began making more appearances as Doof's lovable but bungling henchman, and even had two episodes centered around him ("A Real Boy" & "Norm Unleashed").
Of the Fireside Girls, Ginger has had noticeably more focus than the others starting in season 4, probably because her relationships with Stacy and Baljeet provide additional dynamics beyond simply being backup support for the title characters.
In 1932, E.C. Segar created a character named "Bluto the Terrible" to serve as Popeye's adversary for a single storyline in the Thimble Theatre strip. After the story ended, Segar discarded the character and never used him again... but when Popeye was adapted to animation a year later, Bluto became the series' main villain.
Rumor has it Segar created Bluto at the request of Fleischer Studios so that they'd have a recurring villain to use in the cartoons. And evidently, when creating him, Segar decided he may as well do a story with the character.
Popeye himself is perhaps the greatest example of this, actually. The comic strips he starred in began with chronicling the adventures of the Oyl family. Popeye was going to be a one arc gag, but became so popular that he ascended to the main character of the franchise..
Cornchip Girl from Recess started out as a nameless background character. Around season four or five, she became a more prominent member of the cast, and became the unnoficial seventh member of the gang.
Miss Grotke, the gang's fourth grade teacher, was only in a few episodes of the first season. Come season two and she's in almost every episode of the season. Though by season five, she got Demoted to Extra and suffered from Chuck Cunningham Syndrome in the final season.
CJ also applies. She began as a character Mordecai spent time with in Season 3's "Yes Dude Yes" when he mistakenly thought his love interest Margaret had become engaged to another guy. After Margeret left to go to college at the end of Season 4, CJ was reintroduced and has become both Mordecai's love interest and Eileen's friend.
Filburt the turtle was initially a minor recurring character in Rocko's Modern Life who served as a nerdy employee for various jobs. He would go on to become one of Rocko's friends and have larger roles later on in the cartoon.
Others notable Simpsons promotions include Disco Stu, the Sea Captain, Hans Moleman, and many others. Disco Stu is a particularly obvious case, as he was only introduced for the sake of a single joke, but still pops up every so often.
Apu Nahasapeemapetilon started off as a character whose only role was being the local convenience store owner, with little characterisation beyond that. It stayed like this for a bit, being little more than an extra, but soon changed. He's now become one of the most iconic secondary characters, receiving several focus episodes, becoming the only non-Simpsons player character in The Simpsons: Hit & Run, and becoming far more rounded and deep.
Bart's best friend Milhouse wasn't even an extra on the show—he was first seen as an unnamed kid on a Simpsons "Butterfinger" commercial.
Apparently, though not exactly an extra, Tails was also due to get a bigger role in S3 as well (he was rather low-key in the first [and only] two seasons).
South Park has many instances of this, and the show is fairly notable for promoting far more background characters into prominence than introducing new ones altogether. Virtually every member of the boys' class now has an established personality to some degree.
Butters in particular started off as a Living Prop before being increasingly used as a supporting background character in seasons 3-5 and eventually becoming the Fifth Ranger. He is now pretty much a main character with more focus than Kenny. This happened around the same time Kenny was more or less Demoted to Extra, though it's hard to say if Butters' rise is the cause or effect of that. Maybe it's just that Butters is capable of talking.
The next South Park game coming out features him lined up with the other four boys on the cover, confirming his place as a member of the gang.
Randy Marsh started out being the role of Stan's dad and geologist before being promoted the most prominent adult character with many episodes dedicated to him whenever he does something stupid.
Karen in Spongebob Squarepants. The more recent the episode starring Plankton, the more likely Karen will play an important role.
Speaking of Plankton, he has been appearing more in the later seasons compare to the early seasons.
Hotstreak only appeared in several issues of the original Static comics, and was defeated pretty easily due to his Weaksauce Weakness. Static Shock removed said weakness and gave him a much larger role, effectively making him Static's Arch-Enemy.
Several minor one-off villains from the comics like Rubberband Man and Puff also had much larger roles in the cartoon than they ever did in the source material.
The inmates in Superjail! were mostly interchangeable minor characters, aside from a few that were given distinct designs or personalities. With the second season, some of these inmates had their roles greatly expanded, speaking more often and having more to do with the plots. The third season continued this sort of development. The creators have stated that if they come to like a particular inmate design, they'll do what they can to reuse them.
Team Umizoomi's Umi Car started out as just transportation for the team that eventually disappears for the rest of the story. Midway through season 1, he gained a face but still didn't ascend from his role as an extra until season 2, where he has gotten several major roles.
Leatherhead in Teenage Mutant Ninja Turtles (2012). After being a guest character for the majority of the first and second seasons, he takes on a pivotal role in the second season finale and becomes a recurring character in the third.
The "Two-Tone Town" episode of Tiny Toon Adventures reintroduced a trio of obscure Merrie Melodies characters from the black-and-white 1930s shorts: Foxy and Roxy (obvious Mickey and Minnie Mouse ripoffs) and Goopy Geer (who would be an obvious Goofy ripoff if he didn't predate Goofy). They were modernized and given more distinct personalities for their Tiny Toons appearance; most viewers were probably unaware that they were actual characters from the black-and-white era and not just Retraux imitations like the subsequent Warner siblings.
Also, Gogo Dodo and his residence of Wackyland are a large part of the show, to the point that Gogo appears in the opening theme. He is based on a character from a single 1938 Porky Pig short, not counting the color remake.
Likewise Pumyra was a secondary character the writers of the old show just didn't know what to do with most of the time. In the remake she's confirmed to have an expanded role and even has multiple episodes dedicated to her development.
In the original U.S. Acres comic strip, Orson's nasty brothers only briefly appeared in the first three weeks, and they didn't have names then. Garfield and Friends turned them into recurring villains and gave them the names Mort, Gort, and Wart.
Similarly, Binky the Clown only appeared and/or was referenced in a handful of Garfield comics, but was given a much larger role in the cartoon show.
21 and 24 from The Venture Bros. were minor characters early on in the first season, but became a bit more important as the season went on, and then starting with season 2 they were part of the main cast. Same could probably be said of the Monarch who went from a recurring character to getting equal screen time to the Venture family.
21 and 24's ascension is actually lampshaded a couple times, once by 21 himself ("We're like main characters!") and later by the Monarch ("They have that weird mix of expendable and invulnerable that makes for a perfect henchman."). Later on in the latter episode, 21 and 24 reiterate their status to a new character, claiming he's the Red Shirt on their mission.
The creators often mention looking over crowd scenes and attributing characterization to whoever catches their eye, many showing up later in the show as actual characters.
Such as Sgt. Hatred. He was initially a throwaway off-screen villain that the 21 & 24 stole equipment from, to being one of the many unnamed villains who became excited at Dr. Venture's Walking Eye in Season 2, he eventually got a big plot upgrade in Season 3 where he eventually became Venture's new Guild arch and became tied to the backstory of why Brock was assigned to the Venture family. As of Season 4, he's become one of the main characters, becoming the new Venture bodyguard!
Elyon in the animated version of W.I.T.C.H. is portrayed this way. Those only familiar with the show may or may not be aware that in the original comics, Elyon was a major character from the beginning. However, the TV series deliberately portrays her as a seemingly minor character who gradually "ascends" to being a major character .
Truth be told, a lot of characters in WITCH are ascended extras, the biggest ones being the previous Guardians minus Nerissa - in the comics, Yan Lin dies halfway through the second issue and becomes a member of the Council, Halinor's been dead, Cassidy was just an exposition ghost and Kadma was the bitter old lady. In the cartoon, Yan Lin's alive and The Mentor, Halinor's alive and part of the Council, Cassidy actually comes back to life and Kadma becomes a ruler of another world.
Woody Woodpecker's girlfriend Winnie Woodpecker was only shown in one classic Woody Woodpecker cartoon called Real Gone Woody, but she was mainly shown in comic books as a recurring character. It wasn't until later in The New Woody Woodpecker Show that she became an active member of the series, with a personality similar in silliness to Woody, though showing a more developed sense of dignity.
Some one-shot villains on WordGirl might qualify. For example, Ms. Question, a villain previously seen as a student of the Coach, (another villain), has recently had two episodes where she was the main villain (after gaining the power to confuse people), as has smooth-talking game show host Seymour Orlando Smooth, who had previously only appeared in one episode. In fact, the only WordGirl villain who hasn't returned at least once is the Masked Meat Marauder (although he's probably enjoying his time in that other city with lots of butcher shops and no superhero girls or monkey sidekicks).
Artemis and her family in Young Justice. Artemis is based on a D-list (or lower) character in the comics while she is part of a team of young superheroes like Robin, Superboy and Kid Flash for the show. Her supervillain parents and sister become more prominent because of this as well. Said sister isn't even related to her in the comics where she's the most prominent of the lot, but thanks to the writers mixing up DC Comics lore she gets an even larger role as one of the heroes' more prominent adversaries.
The second season premiere, "Happy New Year," is a long parade of Ascended Extras: Bumblebee is one of M'gann's and Conner's friends from high school, Lagoon Boy had a single-line cameo when Kaldur visited his old class in "Downtime," Beast Boy was a character of the day in "Image," and Batgirl had a single-line cameo in "Home Front," plus silent cameos in "Failsafe" and "Misplaced." Mal Duncan actually ascended twice: he's promoted from Conner and M'gann's background high school friend to the team's Mission Control, then from Mission Control to Badass Normal hero.
In the Nickelodeon series Peter Rabbit, Cottentail is a main character and even has a major role in a few episodes.
Mittens also becomes a recurring in the series despite her giving the least focus in The Tale of Tom Kitten and her mother and her the other siblings (Most notable Tom Kitten) aren't seen in the series itself.