Thymopoietins (TMPOs, previously abbreviated TPs) α (75 kDa), β (51 kDa), and γ (39 kDa) are related nuclear proteins expressed in many or all tissues. TMPO α is present diffusely throughout the nucleus, while TMPOs β and γ are localized to the nuclear membrane. Here we report the cloning and analysis of a single TMPO gene encoding TMPOs α, β, and γ, which are produced by alternative mRNA splicing, as previously inferred from cDNA sequences. The eight exons of the TMPO gene are spread over ∼35 kb. Exon 4, which is spliced into TMPO α mRNA, contains sequences that encode a putative basic nuclear localization motif. Exon 8, which is spliced into TMPO β and γ mRNAs, encodes a hydrophobic putative membranespanning domain that is thought to target TMPOs β and γ to the nuclear membrane. TMPO β appears to be the human homologue of the recently described rat protein LAP2 (lamina-associated polypeptide 2), which is thought to play an important role in the regulation of nuclear architecture by binding lamin B1 and chromosomes in a manner regulated by phosphorylation during mitosis (K. Furukawa and L. Gerace, La Jolla, pers. comm., 22 Nov. 1994). The human TMPO gene maps to chromosome band 12q22.