Fast skeletal muscle myosin light chains 1 and 3 are produced from a single gene by a combined process of differential RNA transcription and splicing

Fast myosin light chains (MLC(f)) 1 and 3 are proteins associated with the thick filament in vertebrate fast muscle fibers. MLC1(f) and MLC3(f) have complete sequence homology for the first 141 amino acids from their COO^- end, but they differ in length and amino acid sequence at their NH₃⁺ ends (MlC1(f) = 49 amino acids, MLC3(f) = 8 amino acids), and they are translated from different mRNAs. To elucidate the structural relationship between mammalian fast myosin light chain 1 and 3 isoforms, cDNA clones have been isolated from rat skeletal muscle and their primary nucleotide sequences were determined. MLC1(f) and MLC3(f) mRNAs display complete sequence identity in the 3' untranslated sequences (285 base pairs) and in the codons specifying the 142 carboxyl-terminal amino acids. In contrast, the sequences encoding the amino-terminal 49 and 8 amino acids of MLC1(f) and MLC3(f), respectively, as well as their 5' untranslated regions, are highly divergent. Using the cloned cDNAs as probes we have isolated the single gene locus encoding both MLC1(f) and MLC3(f) in four overlapping genomic clones spanning over ~25 kilobase pairs (kb) of DNA. The sequences encoding the common body of MLC1(f) and MLC3(f) are distributed in 4 separate exons at the 3' end of the gene. The start of transcription site and 5' untranslated region of MLC3(f) is found 5 kb upstream from the common body while the MLC1(f) mRNA transcription start site and the exon specifying the 5' untranslated region and amino acid 1-40 of MCL1(f) lies about 10 kb further upstream. Strikingly, a mini-exon that specifies amino acids 41-49 of MLC1(f) is located downstream from the two MLC3(f)-specific exons (5' untranslated and amino acids 1-8) and is separated from the upstream MLC1(f) exons by 12 kb of DNA. This bizarre gene organization implies a novel form of alternative promoter utilization and RNA splicing that is tissue-specific and developmentally regulated in order to generate the mature MLC1(f) and MLC3(f) mRNAs from a single gene.