We have isolated and analyzed the structure of the genes coding for the alpha and beta forms of the human cardiac myosin heavy chain (MYHC). Detailed analysis of four overlapping MYHC genomic clones shows that the alpha-MYHC and beta-MYHC genes constitute a total length of 51 kilobases and are tandemly linked. The beta-MYHC-encoding gene, predominantly expressed in the normal human ventricle and also in slow-twitch skeletal muscle, is located 4.5 kilobases upstream of the alpha-MYHC-encoding gene, which is predominantly expressed in normal human atrium. We have determined the nucleotide sequences of the beta form of the MYHC gene, which is 100% homologous to the cardiac MYHC cDNA clone (pHMC3). It is unlikely that the divergence of a few nucleotide sequences from the cardiac beta-MYHC cDNA clone (pHMC3) reported in a MYHC cDNA clone (pSMHCZ) from skeletal muscle is due to a splicing mechanism. This finding suggests that the same beta form of the cardiac MYHC gene is expressed in both ventricular and slow-twitch skeletal muscle. The promoter regions of both alpha- and beta-MYHC genes, as well as the first four coding regions in the respective genes, have also been sequenced. The sequences in the 5'-flanking region of the alpha- and beta-MYHC-encoding genes diverge extensively from one another, suggesting that expression of the alpha- and beta-MYHC genes is independently regulated.