Abstract: Previous studies have shown that the distribution of genes in prokaryotes and eukaryotic genomes is not random. Using the thousands of cellular functions that appear in the Gene Ontology (GO) project, we exhaustively studied the relation between functionality and genomic localization of genes across 16 organisms with rich GO ontologies (one prokaryote and 15 eukaryotes). Overall, we found that the genomic distribution of cellular functions tends to be more similar in organisms that have higher evolutionary proximity. At the primary level, which measures localization of functionally related genes, the prokaryote Escherichia coli exhibits the highest level of organization, as one would expect given its operon-based genomic organization. However, examining a higher level of genomic organization by analyzing the co-localization of pairs of different functional gene groups, we surprisingly find that the eukaryote yeast Saccharomyces cerevisiae is markedly more organized than E. coli. A network-based analysis further supports this notion and suggests that the eukaryotic genomic architecture is more organized than previously thought. See online Supplementary Material at (www.liebertonline.com).