The runt-related transcription factor 3 (RUNX3) gene is located on chromosome 1p36.11. Its product is a known regulator of major developmental pathways. The gene encodes a transcription factor of 415 amino acids with approximately 44 kDa molecular weight. It is a member of the runt domain-containing family of transcription factors that function as tumor suppressors, and interacts with other transcription factors. It can either activate or suppress transcription by forming a complex of a heterodimer of RUNX3 protein and a beta subunit which binds to the core DNA sequence 5'-PYGPYGGT-3' that is found in a number of enhancers and promoters.

The RUNX3 gene is composed of five coding exons and spans about 66 kb in size. It has multiple transcript variants encoding different isoforms. It has been found that this gene is frequently deleted or transcriptionally silenced in solid tumors, including pancreatic, bladder, gastric, colorectal, lung, and others. This inactivation is due to DNA hypermethylation.