Structure and Expression of the Murine Slow/Cardiac Troponin C Gene

Abstract

Cardiac troponin C (cTnC) is the calcium-binding subunit of the myofibrillar thin filament that regulates excitation-contraction coupling in cardiac muscle. We have utilized a novel polymerase chain reaction cloning procedure to isolate cDNA clones encoding murine cTnC. Murine cTnC is a 161-amino acid polypeptide that has been highly conserved during evolution. Southern blot analyses demonstrated that the cTnC gene is a member of a multigene family. Northern blot analyses revealed that the cTnC gene is expressed in murine cardiac tissue and slow skeletal muscle (soleus), but is not expressed in fast skeletal muscle (extensor digitorum longus and anterior tibialis) or in neonatal or adult brain, kidney, lung, liver, or testis. In addition, while the cTnC gene is not expressed in murine C2C12 myoblasts, differentiation of these cells into myotubes was shown to result in a dramatic induction of cTnC gene expression. A full length cTnC genomic clone was isolated from a murine genomic library by hybridization with a cTnC cDNA probe and structurally characterized by DNA sequence, primer extension, and S1 nuclease protection analyses. The cTnC gene is 3.4 kilobase pairs long and is composed of six exons. The introns do not appear to divide the gene into functional domains. Analysis of the 5'-flanking region of the gene revealed the presence of a consensus TATA box 24 base pairs 5' of the transcription start site. Despite the finding that the gene is expressed only in cardiac and slow skeletal muscle, it lacks the previously described CArG and M-CAT transcriptional regulatory sequence motifs that are involved in regulating the expression of a number of other myofibrillar genes.