Alpha 1-adrenergic receptor stimulation of sarcomeric actin isogene transcription in hypertrophy of cultured rat heart muscle cells.

During pressure-load hypertrophy of the adult heart in vivo, there is up-regulation of the mRNA encoding skeletal alpha-actin, the sarcomeric actin iso-mRNA characteristic of mature skeletal muscle and the fetal/neonatal heart. We have shown previously that during alpha 1-adrenergic receptor-stimulated hypertrophy of cultured rat heart myocytes, the induction of skeletal alpha-actin mRNA is greater than that of the mRNA encoding cardiac alpha-actin, the sarcomeric actin iso-mRNA characteristic of the adult heart. To determine if this actin iso-mRNA switch during cardiac hypertrophy reflects changes in the transcriptional status of the myocyte nucleus, we quantified the rate of transcription of actin mRNAs and total RNA, using an in vitro run-on transcription assay with nuclei isolated from the cultured myocytes after stimulation with norepinephrine (NE). Transcription of skeletal alpha-actin was increased at 3 h after NE, reached a maximum 6.1-fold increase at 12 h, and returned to the control level at 24 h. The EC50 for NE was 200 nM, and pharmacologic studies indicated alpha 1-receptor specificity. Transcription of cardiac alpha-actin was also increased rapidly by NE (maximum 4.6-fold vs. control at 3 h). However, cardiac alpha-actin transcription had returned to the control level at 6 h, when NE-stimulated skeletal alpha-actin transcription was still increasing. Transcription of the cytoskeletal (beta) actin gene was not changed significantly by NE treatment. Total RNA transcription was not increased until 6 h after NE (1.5-fold vs. control) and remained elevated through 24 h. Inhibition of protein synthesis did not attenuate NE-stimulated actin gene transcription. Thus the alpha 1-adrenoceptor mediates a rapid, transient, and selective increase in transcription of the sarcomeric actin isogenes during cardiac myocyte hypertrophy. Skeletal alpha-actin, the fetal/neonatal isogene, is induced preferentially to cardiac alpha-actin, the adult isogene. The different kinetics of actin isogene and total RNA transcription and the independence of transcription from protein synthesis suggest that transcriptional induction via the alpha 1 receptor is complex and may involve preexisting regulatory factors. These results are the first to demonstrate that the alpha 1-adrenergic receptor is a molecular mediator of transcriptional changes underlying an isogene switch that is known to be associated with cardiac myocyte hypertrophy.

Interleukin-1 beta induces cardiac myocyte growth but inhibits cardiac fibroblast proliferation in culture.

Interleukin-1 (IL-1), initially called "endogenous pyrogen," is primarily known as a mediator of inflammation. However, it also plays many other diverse physiologic roles including the stimulation and inhibition of both primary cells in culture and the interstitial and parenchymal cells of a number of organs including the heart. In the heart, IL-1 expression has traditionally been reported in situations where there is immunologic myocardial injury such as occurs during transplant rejection and congestive heart failure. For this reason, all of the effects of IL-1 have been presumed to be deleterious. Using a cell culture model which allows both the muscle cells (myocytes) and nonmuscle cells (fibroblasts) to be evaluated separately, we have found that IL-1 induces both cardiac myocyte hypertrophy and reinitiates myocyte DNA synthesis. In stark contrast, IL-1 exerts a potent anti-proliferative effect on cardiac fibroblasts. To our knowledge this is the first report concerning the differential effects of IL-1 on myocardial cell growth in culture and, given the inducible expression of IL-1 by myocardial cells during stress, underscores the importance of investigating the complex nature of the intracardiac cell-cell interactions that occur in the heart.

Regulation of thyroid hormone receptor isoforms in physiological and pathological cardiac hypertrophy.

Physiological and pathological cardiac hypertrophy have directionally opposite changes in transcription of thyroid hormone (TH)-responsive genes, including alpha- and beta-myosin heavy chain (MyHC) and sarcoplasmic reticulum Ca(2+)-ATPase (SERCA), and TH treatment can reverse molecular and functional abnormalities in pathological hypertrophy, such as pressure overload. These findings suggest relative hypothyroidism in pathological hypertrophy, but serum levels of TH are usually normal. We studied the regulation of TH receptors (TRs) beta1, alpha1, and alpha2 in pathological and physiological rat cardiac hypertrophy models with hypothyroid- and hyperthyroid-like changes in the TH target genes, alpha- and beta-MyHC and SERCA. All 3 TR subtypes in myocytes were downregulated in 2 hypertrophy models with a hypothyroid-like mRNA phenotype, phenylephrine in culture and pressure overload in vivo. Myocyte TRbeta1 was upregulated in models with a hyperthyroid-like phenotype, TH (triiodothyronine, T3), in culture and exercise in vivo. In myocyte culture, TR overexpression, or excess T3, reversed the effects of phenylephrine on TH-responsive mRNAs and promoters. In addition, TR cotransfection and treatment with the TRbeta1-selective agonist GC-1 suggested different functional coupling of the TR isoforms, TRbeta1 to transcription of beta-MyHC, SERCA, and TRbeta1, and TRalpha1 to alpha-MyHC transcription and increased myocyte size. We conclude that TR isoforms have distinct regulation and function in rat cardiac myocytes. Changes in myocyte TR levels can explain in part the characteristic molecular phenotypes in physiological and pathological cardiac hypertrophy.

The cell-stretcher: A novel device for the mechanical stimulation of cell populations.

Mechanical stimulation appears to be a critical modulator for many aspects of biology, both of living tissue and cells. The cell-stretcher, a novel device for the mechanical uniaxial stimulation of populations of cells, is described. The system is based on a variable stroke cam-lever-tappet mechanism which allows the delivery of cyclic stimuli with frequencies of up to 10 Hz and deformation between 1% and 20%. The kinematics is presented and a simulation of the dynamics of the system is shown, in order to compute the contact forces in the mechanism. The cells, following cultivation and preparation, are plated on an ad hoc polydimethylsiloxane membrane which is then loaded on the clamps of the cell-stretcher via force-adjustable magnetic couplings. In order to show the viability of the experimentation and biocompatibility of the cell-stretcher, a set of two in vitro tests were performed. Human epithelial carcinoma cell line A431 and Adult Mouse Ventricular Fibroblasts (AMVFs) from a dual reporter mouse were subject to 0.5 Hz, 24 h cyclic stretching at 15% strain, and to 48 h stimulation at 0.5 Hz and 15% strain, respectively. Visual analysis was performed on A431, showing definite morphological changes in the form of cellular extroflections in the direction of stimulation compared to an unstimulated control. A cytometric analysis was performed on the AMVF population. Results show a post-stimulation live-dead ratio deviance of less than 6% compared to control, which proves that the environment created by the cell-stretcher is suitable for in vitro experimentation.

Signaling pathways responsible for fetal gene induction in the failing human heart: evidence for altered thyroid hormone receptor gene expression.

BACKGROUND: We have previously demonstrated that changes in myosin heavy chain (MHC) isoforms that occur in failing human hearts resemble the pattern produced in rodent myocardium in response to hypothyroidism. Because thyroid hormone status is usually within normal limits in these patients, we hypothesized that failing/hypertrophied human myocardium might have a defect in thyroid hormone signaling due to alterations in expression of thyroid hormone receptors (TRs). METHODS AND RESULTS: To examine this hypothesis, we used RNase protection assay to measure mRNA levels of TRs in failing left ventricles that exhibited a fetal pattern of gene expression, ie, decreased expression of alpha-MHC with increased beta-MHC expression compared with left ventricles from age-matched controls. We detected expression of TR-alpha(1), -alpha(2), and -beta(1) isoforms in human left ventricles. In failing left ventricles, TR-alpha(1) was downregulated, whereas TR-alpha(2), a splice variant that does not bind thyroid hormone but inhibits responses to liganded TRs, was increased. Expression levels of TR-beta(1) did not differ significantly between the 2 groups. According to linear regression analysis, expression levels of TR-alpha(1) and -alpha(2) were positively and negatively correlated with those of alpha-MHC, respectively. CONCLUSIONS: We conclude that decreases in TR-alpha(1) and increases in TR-alpha(2) may lead to local attenuation of thyroid hormone signaling in the failing human heart and that the resulting tissue-specific hypothyroidism is a candidate for the molecular mechanism that induces fetal gene expression in the failing human ventricle.

Autocrine and Paracrine Regulation of Myocardial Cell Growth in Vitro The TGFß Paradigm.

Over the past few years, there has been an increasing appreciation of the role that growth factors/cytokines play in cardiac growth/development and the myocardial response to injury. This has derived, at least in part, from two observations: (a) that a number of these factors are expressed in response to myocardial stress, and (b) that some of these factors can stimulate myocardial growth in culture along with the characteristic set of gene products that are associated with hypertrophy in vivo. Virtually all of the cells that make up the adult myocardium have at one time or another been reported to either be the source, the site of action, or both, of many of these cytokines. Although the cell specificity of cytokine production is not critical to the understanding of the complex nature of intracardiac cell-cell interactions as a mechanism of cardiac growth/development, this distinction does have some importance as myocardial cell culture gains increased popularity in the research community. Because there are no true cardiac muscle cell lines, all of the myocyte cultures used in these studies are, in essence, "cocultures" of cardiac myocytes and nonmyocytes (predominantly fibroblasts), with absolute numbers of these "contaminating" cells a function of plated cell density. As such, the cell specificity of some substances on myocardial cell growth and transcriptional program (of both myocytes and fibroblasts) is not always clear. This makes the critical examination of the expression (and effects) of these growth-promoting substances in culture of particular importance with attention to the specific culture conditions employed. The complexity of the situation in the heart is compounded by the observation that many of these substances have differential effects on the individual myocardial cell types and can induce the expression of other cytokines/cytokine receptors by these cells. In this report, the investigations published to date on the autocrine/paracrine effects of these factors on myocardial cell growth in culture are reviewed. The complexity of the subject is illustrated with the findings from our laboratory investigating one of the cytokines with growth-promoting effects on myocardial cells in culture, transforming growth factor beta (TGFß). © 1996, Elsevier Science Inc. (Trends Cardiovasc Med 1996;6:217-226).

Angiotensin II stimulates cardiac myocyte hypertrophy via paracrine release of TGF-beta 1 and endothelin-1 from fibroblasts.

OBJECTIVE: We sought to determine whether angiotensin II (Ang II) promotes hypertrophy of cardiac directly or via paracrine mechanisms mediated by cardiac fibroblasts. METHODS: We studied neonatal rat cardiac myocytes and fibroblasts in culture as a model system. Paracrine effects of Ang II were identified using conditioned medium and co-culture experiments. RESULTS: Ang II type 1 (AT1) receptors responsible for myocyte growth localized to fibroblasts in radioligand binding, emulsion autoradiography, Western analysis, and immunofluorescence staining experiments. The bulk of AT1 receptor binding in myocyte cultures (1343 +/- 472 sites/cell) was to Ang II receptors on contaminating fibroblasts (9747 +/- 2126 sites/cell). Ang II induced significant paracrine trophic effects on myocytes in conditioned medium (40% increase in protein synthesis over control) and co-culture (4-fold increase over control) experiments. TGF-beta 1 and endothelin-1 were paracrine mediators of hypertrophy in neutralization experiments. CONCLUSIONS: Ang II stimulates cardiac myocyte hypertrophy via paracrine release of TGF-beta 1 and endothelin-1 from cardiac fibroblasts in a neonatal rat cell culture model.

Hypoxia differentially regulates stress proteins in cultured cardiomyocytes: role of the p38 stress-activated kinase signaling cascade, and relation to cytoprotection.

OBJECTIVE: Stress proteins (heat shock proteins, HSPs) are molecular chaperones that have been shown to enhance the survival of cells exposed to environmental stress. We sought to investigate the effects of hypoxia on the levels of HSP27 and heme oxygenase-1 (HO-1 or HSP32) in an established model of rat neonatal cardiac myocytes in culture. METHODS: Myocytes were subjected to hypoxia (<0.5% O(2) for 16 h). Studies of cell viability and nuclear morphology showed no evidence of cell death under these conditions. RESULTS: Messenger RNA analysis demonstrated constitutive expression of HSP27 and low levels of HO-1. Hypoxia strongly induced HO-1 mRNA without affecting HSP27 mRNA. In parallel to mRNA levels, hypoxia increased HO-1 protein level without affecting HSP27. To further assess the signaling pathways implicated in HO-1 induction, we used inhibition experiments. The tyrosine kinase inhibitor tyrphostin and the mitogen-activated protein kinase inhibitor PD98059 did not prevent HO-1 induction, while the protein kinase C inhibitor chelerythrine partially blocked this response. The p38 stress-activated kinase inhibitor SB203580 was the most potent in suppressing hypoxia-induced HO-1. In vitro kinase assays, cell labeling and immunoprecipitation showed activation of signaling pathways downstream of p38 stress-activated kinase as revealed by an increase in phosphorylation of MAPKAPK-2/3 kinases and HSP27. CONCLUSIONS: These data show a differential pattern of hypoxia-induced HSP expression and implicate the stress kinase in HO-1 induction. Thus, selective regulation of HSP levels may play a role in the cardioprotective mechanisms that participate in the adaptive response to hypoxia-induced stress.

Analysis of long- and short-range contribution to adhesion work in cardiac fibroblasts: an atomic force microscopy study.

Atomic force microscopy (AFM) for single-cell force spectroscopy (SCFS) and Poisson statistic were used to analyze the detachment work recorded during the removal of gold-covered microspheres from cardiac fibroblasts. The effect of Cytochalasin D, a disruptor of the actin cytoskeleton, on cell adhesion was also tested. The adhesion work was assessed using a Poisson analysis also derived from single-cell force spectroscopy retracting curves. The use of Poisson analysis to get adhesion work from AFM curves is quite a novel method, and in this case, proved to be effective to study the short-range and long-range contributions to the adhesion work. This method avoids the difficult identification of minor peaks in the AFM retracting curves by creating what can be considered an average adhesion work. Even though the effect of actin depolymerisation is well documented, its use revealed that control cardiac fibroblasts (CT) exhibit a work of adhesion at least 5 times higher than that of the Cytochalasin treated cells. However, our results indicate that in both cells short-range and long-range contributions to the adhesion work are nearly equal and the same heterogeneity index describes both cells. Therefore, we infer that the different adhesion behaviors might be explained by the presence of fewer membrane adhesion molecules available at the AFM tip-cell interface under circumstances where the actin cytoskeleton has been disrupted.

Kinetics of carnitine-dependent fatty acid oxidation: implications for human carnitine deficiency.

The relationship between the concentration of carnitine and the oxidation of oleate was examined in homogenates prepared from skeletal muscle, liver, kidney, and heart of the rat, and from canine and human skeletal muscle. the carnitine content of these tissues in situ spanned a wide range, from about 0.1 mumol per gram in rat liver to about 3.0 mumol per gram in human muscle. The concentration of carnitine required for half-maximal rates of fatty acid oxidation in vitro also varied greatly (10 to 15 microM for rat liver to 200 to 400 microM for human muscle), and in rough proportion to the normal carnitine content of the tissues. For any given tissue, the carnitine content seems to be set at a level necessary for optimal rates of fatty acid oxidation. The data provide a plausible explanation for the fact that muscle fatty acid metabolism is severely impaired in the syndrome of human carnitine deficiency, since measured carnitine levels are in the range expected to limit substantially the capacity for fatty acid oxidation.

Expression of skeletal muscle sarcoplasmic reticulum calcium-ATPase is reduced in rats with postinfarction heart failure.

OBJECTIVE: To determine whether heart failure in rats is associated with altered expression of the skeletal muscle sarcoplasmic reticulum Ca2+-ATPase (SERCA). METHODS: SERCA protein and mRNA were examined in the soleus muscles of eight female rats with heart failure induced by coronary artery ligation, six weeks after the procedure (mean (SEM) left ventricular end diastolic pressure 20.4 (2.2) mm Hg) and in six sham operated controls by western and northern analyses, respectively. RESULTS: SERCA-2a isoform protein was reduced by 16% (112 000 (4000) v 134 000 (2000) arbitrary units, p < 0.001), and SERCA-2a messenger RNA was reduced by 59% (0.24 (0. 06) v 0.58 (0.02) arbitrary units, p < 0.001). Although rats with heart failure had smaller muscles (0.54 mg/g v 0.66 mg/g body weight), no difference in locomotor activity was observed. CONCLUSIONS: These results may explain the previously documented abnormalities in calcium handling in skeletal muscle from animals with the same model of congestive heart failure, and could be responsible for the accelerated muscle fatigue characteristic of patients with heart failure.

The Ultraviolet Index: a useful tool.

The Ultraviolet Index was developed in the United States in 1994 following successful use of ultraviolet (UV) alerts in other countries. This daily National Weather Service prediction is a calculation which integrates five data elements to yield the amount of UV radiation impacting the surface (1m2) at solar noon in 58 of the largest US population centers. This simple numeric prediction is then categorized by the Environmental Protection Agency into five "exposure levels" with protective actions recommended for each level. This information is disseminated through the media. Daily reminders seem to affect awareness and behavior in Canada, but US surveys indicate the need for better understanding through educational graphics. Comparing the UV Index to a precipitation prediction has merit in that it links a familiar daily prediction with implied appropriate protective measures. Graphics link the ideas that "when it rains it pours and when it shines it radiates." Beginning in schools, camps, and dermatology meetings, using the rain/shine analogy, a wider exposure to the Ultraviolet Index is proposed.

Trophic factors for cardiac myocytes.

During early neonatal myocardial growth, cardiac myocytes undergo a terminal differentiation stage after which cellular proliferation no longer occurs. Subsequent growth occurs by means of enlargement/hypertrophy of the existing cells. In an effort to dissect the molecular mechanisms underlying myocardial hypertrophy, a cell culture model of cardiac hypertrophy was developed in our laboratory. In this model, neonatal rat heart muscle cells respond to alpha 1-adrenergic receptor stimulation with an increase in cell size, total protein and the cellular contents of several messenger (m)RNA encoding fetal/neonatal contractile protein isoforms, and also that encoding the proto-oncogene c-myc. Similar changes in gene expression are seen in pressure-load hypertrophy in vivo. Recent observations on the production of peptide growth factors by the myocardium suggest an additional role of cell-cell interaction in cardiac growth and development. Some of the growth factors which have been found in normal myocardium include platelet-derived growth factor B chain, fibroblast growth factor (both acidic and basic), transforming growth factor beta, insulin-like growth factor-1 and insulin-like growth factor-2, and nerve growth factor. We recently identified a heparin-binding growth factor produced by cardiac non-myocytes in culture, which acts in a dose-dependent fashion to produce hypertrophy of cardiac muscle cells in culture. Preliminary work suggests that this is dissimilar from previously identified growth factors. Furthermore, the quantitative response of the cardiac myocytes to our growth factor exceeds that seen for other factors. Further work is necessary to ascertain how these factors and the alpha 1-adrenergic system interact to produce the different clinical forms of myocardial hypertrophy.

Exploring the elasticity and adhesion behavior of cardiac fibroblasts by atomic force microscopy indentation.

AFM was used to collect the whole force-deformation cell curves. They provide both the elasticity and adhesion behavior of mouse primary cardiac fibroblasts. To confirm the hypothesis that a link exists between the membrane receptors and the cytoskeletal filaments causing therefore changing in both elasticity and adhesion behavior, actin-destabilizing Cytochalsin D was administrated to the fibroblasts. From immunofluorescence observation and AFM loading/unloading curves, cytoskeletal reorganization as well as a change in the elasticity and adhesion was indeed observed. Elasticity of control fibroblasts is three times higher than that for fibroblasts treated with 0.5 µM Cytochalasin. Moreover, AFM loading-unloading curves clearly show the different mechanical behavior of the two different cells analyzed: (i) for control cells the AFM cantilever rises during the dwell time while cells with Cytochalasin fail to show such an active resistance; (ii) the maximum force to deform control cells is quite higher and as far as adhesion is concern (iii) the maximum separation force, detachment area and the detachment process time are much larger for control compared to the Cytochalasin treated cells. Therefore, alterations in the cytoskeleton suggest that a link must exist between the membrane receptors and the cytoskeletal filaments beneath the cellular surface and inhibition of actin polymerization has effects on the whole cell mechanical behavior as well as adhesion.

The role of interleukin-1 in the failing heart.

The prevalence of congestive heart failure and its continued poor prognosis despite presently available therapeutic options emphasize the importance of pursuing the observations suggesting an important role for an immunomodulatory approach to decompensated cardiac failure. Furthermore, there are several pieces of background information that suggest that cytokines like IL-1 may play a significant role in the pathogenesis of several forms of myocardial dysfunction. Although it seems clear that IL-1 is not acting alone under circumstances of myocardial injury, but in concert with other pro-inflammatory molecules and their effectors, we believe that continued investigations into the cytokine hypothesis will ultimately increase the understanding of how pro-inflammatory molecules influence myocardial function and how the modulation of such factors may improve the myocardial response to injury. The specific observations that emphasize the importance of pursuing a substantive role for IL-1 in this process are: (1) IL-1 is elevated in several cardiac disease states, (2) IL-1 is produced by myocardial cells themselves in response to injury, (3)The alterations in gene expression seen in response IL-1 resembles in many ways the phenotype of the failing heart, and (4) The co-localization of the IL-1 response with that of several previously described negative transcriptional regulators (making them potential targets for therapeutic manipulation).

Transcriptional repression of an embryo-specific muscle gene.

Skeletal muscle involves both the induction and repression of gene expression. Although activation and up-regulation of several contractile protein genes has been shown to occur via transcriptional mechanisms, the mechanisms by which contractile protein genes are repressed during muscle development remain unknown. However, a post-transcriptional mechanism has been implicated in the repression of thymidine kinase expression during muscle development. The chicken cardiac troponin T (cTNT) gene is expressed in early embryonic skeletal muscle but is abruptly repressed in late embryonic/fetal development. Using run-on transcription assays we demonstrate here that cTNT gene repression occurs at the level of transcription. Thus, transcriptional as well as post-transcriptional mechanisms operate both to activate and repress gene expression during skeletal muscle development.

A growth factor for cardiac myocytes is produced by cardiac nonmyocytes.

Cardiac nonmyocytes, primarily fibroblasts, surround cardiac myocytes in vivo. We examined whether nonmyocytes could modulate myocyte growth by production of one or more growth factors. Cardiac myocyte hypertrophic growth was stimulated in cultures with increasing numbers of cardiac nonmyocytes. This effect of nonmyocytes on myocyte size was reproduced by serum-free medium conditioned by the cardiac nonmyocytes. The majority of the nonmyocyte-derived myocyte growth-promoting activity bound to heparin-Sepharose and was eluted with 0.75 M NaCl. Several known polypeptide growth factors found recently in cardiac tissue, namely acidic fibroblast growth factor (aFGF), basic FGF (bFGF), platelet-derived growth factor (PDGF), tumor necrosis factor alpha (TNF alpha), and transforming growth factor beta 1 (TGF beta 1), also caused hypertrophy of cardiac myocytes in a dose-dependent manner. However, the nonmyocyte-derived growth factor (tentatively named NMDGF) could be distinguished from these other growth factors by different heparin-Sepharose binding profiles (TNF alpha, aFGF, bFGF, and TGF beta 1) by neutralizing growth factor-specific antisera (PDGF, TNF alpha, aFGF, bFGF, and TGF beta 1), by the failure of NMDGF to stimulate phosphatidylinositol hydrolysis (PDGF and TGF beta 1), and, finally, by the apparent molecular weight of NMDGF (45-50 kDa). This nonmyocyte-derived heparin-binding growth factor may represent a novel paracrine growth mechanism in myocardium.

Interleukin-1beta is a negative transcriptional regulator of alpha1-adrenergic induced gene expression in cultured cardiac myocytes.

We recently reported that interleukin-1beta (IL-1beta) induces a novel form of cardiac myocyte hypertrophy characterized by an increase in protein content but an absence of the fetal program of skeletal alpha-actin or beta-myosin heavy chain (beta-MHC) gene expression (Palmer, J. N., Hartogensis, W. E., Patten, M., Fortuin, F. D., and Long, C. S. (1995) J. Clin. Invest. 95, 2555-2564). Because of the apparent disparity between this myocardial phenotype and that seen with other hypertrophic agents in culture, such as catecholamines, we investigated the effect of IL-1beta on alpha1-induced cardiomyocyte hypertrophy. Although there was no augmentation in total protein when IL-1beta and phenylephrine were given simultaneously, IL-1beta attenuated the increase in contractile protein mRNAs (skeletal alpha-actin and beta-MHC) in response to phenylephrine. Transient transfection studies with skeletal alpha-actin and beta-MHC promoter constructs linked to the chloramphenicol acetyltransferase (CAT)-reporter gene indicate that repression occurred at the level of gene transcription. In view of the previously reported activity of the zinc finger protein YY1 in the negative regulation of the skeletal alpha-actin promoter in cardiomyocytes (MacLellan, W. R., Lee, T. C., Schwartz, R. J., and Schneider, M. D. (1994) J. Biol. Chem. 269, 16754-16760), we investigated the potential role of this factor in the IL-1beta-mediated effects. Using transient transfection, we found that a mutation in the YY1 binding site of the skeletal alpha-actin promoter abolished the inhibitory effect of IL-1beta. We further found that the 127-base pair fragment of the skeletal alpha-actin promoter required for the IL-1beta effect is also required for inhibition by the overexpression of YY1 in the myocytes. Furthermore, increased levels of YY1 protein are found in IL-1beta treated myocytes. Taken together these results suggest that the repression of contractile protein gene transcription by IL-1beta may be due, at least in part, to activation of the negative transcription factor YY1.