Regulation of troponin T gene expression in chicken fast skeletal muscle: involvement of an M-CAT-like element distinct from the standard M-CAT.

Troponin T (TnT), like other myofibrillar proteins, is expressed as various isoforms in different muscle fibers and/or at different development stages. A recent study suggested the expression pattern of chicken fast TnT isoforms is fixed in a given cell lineage. In the present study, we isolated genomic clones of the chicken fast TnT gene to carry out molecular analysis of its expression mechanism. One of the clones, pWETNTa, contained the 5' upstream region and approximately 20 kb downstream from exon 1. We constructed promoter/upstream segments of the chicken fast TnT gene linked to the bacterial chloramphenicol acetyltransferase (CAT) gene and tested the regulatory function of the 5' upstream region by transient transfection of the gene constructs into muscle cells. We showed that a DNA segment between -264 and -44 bp from the most 5' transcriptional initiation site, which has an MEF2, and M-CAT-like element, a CArG box and two E boxes, was essential for the expression of the fast TnT gene. Furthermore, mutation of the M-CAT-like element in the segment resulted in the most serious reduction in the fast TnT promoter activity. The results suggested that the M-CAT-like element plays an important role in transcriptional regulation of the fast TnT gene. The M-CAT-like element is very similar to the M-CAT element, but in electrophoretic mobility shift assay, the factor(s) that bound to this motif was found to be different from the M-CAT binding factor (MCBF).