The complete DNA sequence is presented of H4-II, the second of the pair of histone H4 genes of the ciliated protozoan, Tetrahymena thermophila. Both H4 genes code for the same protein. Codon usage in these and other Tetrahymena genes is severely restricted and is similar to that in yeast. Flanking regions are AT-rich (greater than or equal to 75%), relative to coding sequences (approximately 45% GC). Except for small, similarly positioned homologies, flanking sequences of the two genes are different. Canonical sequences in higher eukaryotic promoters are not obvious in these genes. Instead, short, localized, base composition eccentricities characterize the 5' flanking sequences of all Tetrahymena genes analyzed. The consensus, P yP u(A)3-4 ATGG initiates translation in these and all other known Tetrahymena genes. Nuclear transcripts and messages of both growing and starved cells begin at multiple sites, mainly at the first or second A residue following a pyrimidine. The palindrome typical of histone message 3' termini in higher organisms is not present. Downstream of both genes are sequences similar to the processing/polyadenylation signal of higher eukaryotes, although the unique 3' ends are not those predicted by the location of the signals.