Body size is an important characteristic for horses of various breeds and essential for the classification of ponies concerning the limit value of 148 cm (58.27 inches) height at the withers. Genome-wide association analyses revealed the highest associated quantitative trait locus for height at the withers on horse chromosome (ECA) 3 upstream of the candidate gene LCORL. Using 214 Hanoverian horses genotyped on the Illumina equine SNP50 BeadChip and 42 different horse breeds across all size ranges, we confirmed the highly associated single nucleotide polymorphism BIEC2-808543 (-log(10)P = 8.3) and the adjacent gene LCORL as the most promising candidate for body size. We investigated the relative expression levels of LCORL and its two neighbouring genes NCAPG and DCAF16 using quantitative real-time PCR (RT-qPCR). We could demonstrate a significant association of the relative LCORL expression levels with the size of the horses and the BIEC2-808543 genotypes within and across horse breeds. In heterozygous C/T-horses expression levels of LCORL were significantly decreased by 40% and in homozygous C/C-horses by 56% relative to the smaller T/T-horses. Bioinformatic analyses indicated that this SNP T>C mutation is disrupting a putative binding site of the transcription factor TFIID which is important for the transcription process of genes involved in skeletal bone development. Thus, our findings suggest that expression levels of LCORL play a key role for body size within and across horse breeds and regulation of the expression of LCORL is associated with genetic variants of BIEC2-808543. This is the first functional study for a body size regulating polymorphism in horses and a further step to unravel the mechanisms for understanding the genetic regulation of body size in horses.