NOTE: This spoiler was sent in by Paul.
The movie starts with a shot of photo-realistic dew building up on a leaf. As the dew drips off the leaf, it sets in motion a round burr from a tree, which rolls menacingly down a hill. It's shot as a huge lumbering thing (not unlike the boulder in "Raiders"). Finally it comes towards a flower, which has a small speck sitting atop it. The burr hits the flower and flings the speck into the air. Throughout the opening credits the speck floats in and out of varying perilous areas until it finally floats past Horton the elephant (Jim Carrey). Horton hears a small voice that seems to be crying for help, but dismisses it as the sounds of his class of young jungle citizens who have gathered for their lesson about Leaf Bugs. The children's parents watch the lesson with elation, praising Horton for his great work as a teacher, however the Sour Kangaroo (Carol Burnett) remains unconvinced, saying that she prefers to "Pouch School" her child, who is never allowed out of his room/pouch.
During the course of the lesson, the speck floats by Horton again and he becomes convinced that there is a small little person holding on to the speck and decides to rescue them. After chasing the speck about he finally finds a small clover to rest the speck on. A few of the jungle citizens are concerned with his behavior, but he doesn't care and tries to get the speck to talk again. While waiting for the speck to speak again, Horton's best friend Morton (Seth Rogan) advises Horton not to tell anyone else that he can hear voices coming from a speck. Horton tries, but is too excited and blurts the information to his students, who promise not to tell anyone unless they promise those people not to tell anyone, which eventually leads to the whole jungle knowing.
In the meantime, we're introduced to the inhabitants of the speck, the citizens of Whoville. A small, but bustling city with a good-hearted, but somewhat absent-minded mayor (Steve Carrell). The mayor spends most of his time giving small increments of advice to his hordes of children, all girls except for one, the quiet and misunderstood son, JoJo (Jesse McCarthy). The mayor loves his son and wants what's best for him and is convinced that the best for him is to become the next in a long line of familial mayors. JoJo feels differently, but doesn't say anything because he's worried about disappointing his dad. In the evening, when his father is busy with the other children, JoJo sneaks off to the abandoned observatory.
The mayor has other concerns also, not the least of which are the constant tremors plaguing the town. The city council feels that it's best to ignore the tremors and berate the mayor in front of the townspeople, explaining that he's just being foolish and that the plans for the town’s centennial celebration will continue unabated. The mayor returns to his office, frustrated at the council’s decision, not to mention the fact that they called him a boob in front of the whole town. While the mayor is grousing in his office, Horton decides to call "hello" very loudly to the speck, rightly assuming that the people may have a hard time hearing him because their ears are so small. His "hello" breaks through the outer boundary of the speck and the sound waves come through a rain pipe outside the mayor’s office.
When the mayor starts talking to the strange voice in the rainpipe, he's convinced it's some sort of prank, however when Horton repeatedly shades the speck and removes the shade, causing rapid succession of day and night, the mayor realizes that his world is indeed on a speck, resting on a clover being held by a giant elephant. The mayor asks Horton to protect his little world and Horton spots a perfect protected area on the top of a mountain. As he journeys to the mountain he imagines the certain adventure he'll undertake in a very imaginative 2-D section spoofing anime-style animation.
Meanwhile, the mayor decides to consult the advice of the smartest professor at Who-University. She tells him that if their world were indeed a small speck, that they would encounter frequent tremors, erratic weather shifts and would possibly be destroyed. This information sends the mayor into panic mode. However, he can't panic for long because he has a dentist appointment.
While he is at the dentist, Horton realizes that the only way to the mountain is across a rickety rope bridge. He walks as gingerly as he possibly can, but the boards start to break anyway. The jerky motion of getting across the bridge is very amusingly spliced with the mayor's dentist visit, which eventually leads to the mayor getting his arm filled with Novocain. He runs home and tells his wife what's happening while trying to remove all potentially dangerous things from his house. She tells him that he's under a lot of stress and that he's entitled to his delusions, but that he shouldn't tell anyone about them.
In the jungle, the Sour Kangaroo decides that she must destroy the speck, as it seems to be a source of contention and potential anarchy. She enlists the help of a vulture named Vlad (Will Arnett) to find Horton and crush the clover. Word of Vlad's impending attack reaches Horton, who in turn warns the mayor that a giant sharp-clawed bird may soon be attacking and almost immediately Vlad's attack happens, causing the town of Whoville to undergo a severe earthquake. When the citizens ask the mayor what's happening he tells them. The city council tries to dismiss his story as nonsense, but the mayor shows the town the rainpipe that he's been talking to Horton through and asks Horton to speak to the town. Horton pauses, trying to think of what to say and Vlad swoops down and takes the clover, leaving the rainpipe in Whoville silent and leaving the mayor without proof of his story.
Horton chases Vlad, but the vulture drops the clover into a giant field of identical clovers, making it next to impossible to find. When the clover hits the ground, the city is severely shaken. Horton runs to the clover field and begins searching through each individual clover. After searching through hundreds of thousands of clovers he spots his clover wafting in the breeze, grabs it and tries to talk to the speck. When there is no answer he fears the worst and we are shown various shots of the debris of Whoville from the crash. However, after a few seconds, the mayor finally answers, the city of Whoville hears Horton and the mayor is proven correct in his story.
Unfortunately, one of the jungle citizens spots Horton talking to the speck again and immediately reports this to the Sour Kangaroo, who decides to rally up the other animals to attack Horton, capture him and destroy the clover. Morton tries to defend Horton, but is spurned, so he rushes to warn his friend.
Morton arrives at the clearing in front of the mountain where Horton is only moments before the mob, but just long enough to let Horton know of their plans. When the citizens arrive, the Sour Kangaroo tells Horton that all he has to do is admit that he was lying and they'll forget the whole thing, however, Horton, being honest and faithful, cannot lie. He tells the mayor that the city must make as much noise as possible so that the other jungle animals will hear them. The town begins screaming "We Are Here!” but the sound is not enough to penetrate the speck. The Wickersham's (the jungle monkeys) grab Horton and begin to cage him. He urges the mayor to have the townspeople scream louder. As the mayor tells everyone to make noise, he sees JoJo running away. Thinking that his son his hiding from the situation, he chases after him. The Sour Kangaroo takes the clover from Horton and holds it above a boiling cauldron of Beezlenut Oil.
The mayor eventually finds JoJo at the observatory, where he discovers that JoJo is a brilliant inventor and musician and has fashioned an enormous musical instrument that uses the concavity of the observatory to blast sound. JoJo fires his machine up and invites his father to watch. The sound is great and bursts through the town, but it's still not enough to get through the speck. When JoJo and the mayor return to the town hall, Horton tells the mayor that it still isn't enough noise. Upon hearing this, JoJo runs to the top of the tallest building in town, just as the Sour Kangaroo drops the clover. JoJo shouts "YAWP!” which is just the small bit of noise needed to finally burst through the outer shell of the speck. When the sound comes out, the Sour Kangaroo's son grabs the clover before it hits the boiling oil. He informs the other jungle citizens that he heard the voices too, everyone becomes quiet and they hear "We are here!" coming from the speck. The baby kangaroo disobeys his mother and returns the clover to Horton as the other citizens allow him passage. The Sour Kangaroo tries to keep them turned against Horton, but they have all heard the Who's and know that Horton was telling the truth.
Horton informs the mayor that they were successful and both worlds celebrate their safety by singing "Can't Fight This Feeling" by REO Speedwagon. (Seriously) Horton notices that the Sour Kangaroo is feeling low because she was disgraced and offers her a chocolate cookie that Vlad the bunny (not to be confused with Vlad the vulture) gave him. She smiles, accepts the offering and joins the group of citizens as they all move up the mountain to bring the Who's to their new protected home.