FSH is a pituitary gonadotropin that is encoded by separate alpha- and beta-subunit genes. We isolated a 12 kilobase (kb) DNA fragment containing the entire human FSH beta gene from a lambda phage genomic DNA library. The nucleotide sequence of the FSH beta gene predicts a 19 amino acid signal sequence and a 111 amino acid apoprotein that differs from the reported protein sequence at three residues and lacks the carboxyterminal eight amino acids, thereby bringing the human FSH beta sequence into register with those described for other mammalian species. Southern blot analyses of human genomic DNA are consistent with a single copy of the FSH beta gene per haploid genome. The FSH beta transcriptional unit spans 3.9 kb and contains two introns. The second intron (1.4 kb) is located between amino acids 35 and 36, a position that is strictly conserved among all of the glycoprotein hormone beta-subunit genes. The first intron occurs 6 base pairs upstream from the start of translation in a location analogous to that of the TSH beta gene. The first exon contains an alternate splicing donor site resulting in 5'-untranslated sequences of 63 (Exon IA) and 33 (Exon IB) bases in length. Approximately 65% of transcripts contain exon IA and 35% contain exon IB. Two different polyadenylation sites are also used. One polyadenylation site coincides with the stop codon, while the other site, which is used in greater than 80% of FSH beta mRNA transcripts, is located approximately 1 kb downstream, resulting in an unusually long 3'-untranslated sequence. The distribution of polyadenylation sites is similar for FSH beta mRNAs containing either Exons IA or IB suggesting that intron processing and polyadenylation are regulated independently. Thus, at least four distinct species of FSH beta mRNA transcripts, all encoding identical peptides, are processed from a single FSH beta gene. The distribution of different FSH beta mRNAs is similar in normal human pituitary tissue and several different FSH producing pituitary adenomas.