Tumor necrosis factor-alpha gene regulation and genetics. Inducer and cell type specific regulation of the human TNF-alpha gene occurs through the recruitment of distinct higher order transcription enhancer complexes, or enhanceosomes. Through a variety of in vitro and in vivo approaches and a cross-species analysis of the TNF-alpha genomic sequence in the primate lineage we show that regulatory regions occur in highly conserved regions of the TNF-alpha locus. We present a primate TNF-alpha gene tree and a map of single nucleotide polymorphisms (SNPs) in the human tumor necrosis factor (TNF)-alpha promoter based upon exploratory sequencing of 333 human TNF-alpha gene promoters from individuals of distinct ancestral backgrounds. We detect ten TNF-alpha promoter SNPs that occur with distinct frequencies in populations of different ancestry and demonstrate that several occur as fixed differences in the primate lineage. These differences are both markers of speciation and subspeciation in the great apes. The correlation of nucleotide sequence variation with TNF-alpha transcriptional activation and genetic polymorphism provides insight into the evolution of non-coding sequences and into how TNF-alpha is controlled in response to different extracellular stimuli.