Thyroid hormone regulates expression of a transfected alpha-myosin heavy-chain fusion gene in fetal heart cells.

Abstract

In ventricular muscle, 3,5,3'-triiodo-L-thyronine (T3) stimulates the expression of the a-myosin heavychain (a-MHC) gene. To test for gene elements required for induction, a fragment of the a-MHC gene containing 2.9 kilobases of 5' flanking sequences and 420 base pairs of DNA 3' to the transcription initiation site was linked to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene. The a-MHC fusion gene was introduced into primary cultures of fetal rat heart myocytes. Induction of the transfected gene was monitored by assaying CAT activity while endogenous a-MHC mRNA expression was measured by using a synthetic oligonucleotide probe complementary to sequences in the 3' untranslated region of the mRNA. Without T3, CAT activity was only slightly greater than background. When T3 at a final concentration of 10 nM was added to the cultures, CAT activity was increased 8-fold by 48 hr. The response time and doses of T3 required for induction of CAT activity and a-MHC mRNA in transfected cells were similar, suggesting that the synthetic and endogenous genes may have a common mechanism of control. When simian virus 40 enhancer and early promoter sequences were included in the construct, CAT activity was constitutively expressed, but it could be increased 7-fold by the addition of T3. Several deletions were introduced into the 5' flanking sequences of the a-MHC fragment and the effects on induction of CAT activity were examined. Progressive deletions of 5' sequences from positions -947 to -374 reduced but did not eliminate induction of CAT activity, suggesting that more than one region may be required for optimal induction by thyroid hormone. The results indicate that DNA sequences required for efficient induction by T3 are present in the 5' flanking sequences of the a-MHC gene.

Fibronectin expression in the normal and hypertrophic rat heart.

“…The changes seen after treatment with triiodothyronine are less likely to be due to fibroblast proliferation and may reflect a response of cardiac myocytes. Triiodothyronine has recently been shown to regulate the expression of myosin isoforms in cultured cardiac myocytes (51) transfected with the 5'-flanking region ofa-myosin heavy chain gene, indicating a direct effect of triiodothyronine on cardiac myocytes. Skeletal a-actin has been shown to be reinduced in cultured rat cardiac myocytes during adrenergic induction of hypertrophy (52).…”

Molecular cloning and characterization of human cardiac alpha- and beta-form myosin heavy chain complementary DNA clones. Regulation of expression during development and pressure overload in human atrium.

“…Significant evidence has been accumulated to suggest that sequences in the 5' flanking regions are important for the regulation oftranscription. Gustafson et al (41) have identified the 5' flanking sequences responsible for the induction by thyroid hormone in the rat a-MHC gene. Furthermore, Saez et al (25) have shown that the 5' flanking sequences of rat and human a-MHC genes are highly homologous.…”

Angiotensin II type 1a receptor gene expression in the heart: AP-1 and GATA-4 participate in the response to pressure overload

“…Plasmids pSV0MCAT and pHSA2000 have been described previously (14,15). The AT 1a R regulatory region was amplified from rat chromosomal DNA using the gene-specific primers 5ЈAT1R and 3ЈAT1R (Table 1).…”