The evolutionary origins and common features of the genes coding for the HMG-14/-17 family of chromosomal proteins have been studied by isolating and sequencing the chicken HMG-14b gene, the true homolog of the human and calf HMG-14 gene. Comparison of the structure of this gene to that of the human HMG-14 gene and to the human and chicken HMG-17 genes indicates that the HMG-14 and HMG-17 genes evolved from a common ancestor. We postulate that the ancestral gene consisted of six exons. In all genes the first exon codes for the entire 5′ untranslated region and for the first four amino acids, which are invariant among all the known members of the HMG-14/-17 protein family. The last exon codes for ten to 16 amino acids and for the entire 3′ untranslated region, which, for each gene, constitutes over 70% of the transcript. The DNA-binding domain of the proteins is encoded by two distinct exons. The genes are characterized by 5′ regions that are highly enriched in G + C residues and have features characteristic of "housekeeping" genes. The HMG-17 genes are distinct from the HMG-14 in that the 5′ regulatory region of the former has two TATA boxes while the HMG-14 genes have no such regulatory element. The chicken HMG-14b gene is a single-copy gene and produces a unique transcript. In this gene, exons II and III are fused and intron 2 is missing. The fusion of the two exons produced a codon for valine in a position that, among all HMG-14/-17 proteins, is unique to HMG-14b. The possible consequences of a valine insertion at the N-terminal end of the DNA-binding domains are discussed. The HMG-14 proteins evolve significantly faster than HMG-17, suggesting that the proteins are subject to different evolutionary pressure. However, certain amino acids are conserved among all the known members of the HMG-14/-17 protein family, suggesting that they are part of the functional domain of this family of chromosomal proteins.