scholarly journals Regulation of calpain and calpastatin in differentiating myoblasts: mRNA levels, protein synthesis and stability

2000 ◽  
Vol 351 (2) ◽  
pp. 413-420 ◽  
Author(s):  
Sivia BARNOY ◽  
Lia SUPINO-ROSIN ◽  
Nechama S. KOSOWER
Keyword(s):  
Transforming Growth Factor ◽  
Protein Translation ◽  
Transforming Growth Factor Β ◽  
Myoblast Fusion ◽  
Myoblast Differentiation ◽  
Mrna Levels ◽  
Endogenous Inhibitor ◽  
Drug Induced ◽  
Dividing Cells ◽  
The Stability

Calpain (Ca2+-dependent intracellular protease)-induced proteolysis has been considered to play a role in myoblast fusion to myotubes. We found previously that calpastatin (the endogenous inhibitor of calpain) diminishes transiently during myoblast differentiation. To gain information about the regulation of calpain and calpastatin in differentiating myoblasts, we evaluated the stability and synthesis of calpain and calpastatin, and measured their mRNA levels in L8 myoblasts. We show here that µ-calpain and m-calpain are stable, long-lived proteins in both dividing and differentiating L8 myoblasts. Calpain is synthesized in differentiating myoblasts, and calpain mRNA levels do not change during differentiation. In contrast, calpastatin (though also a long-lived protein in myoblasts), is less stable in differentiating myoblasts than in the dividing cells, and its synthesis is inhibited upon initiation of differentiation. Inhibition of calpastatin synthesis is followed by a diminution in calpastatin mRNA levels. A similar calpastatin mRNA diminution is observed upon drug-induced inhibition of protein translation. On the other hand, transforming growth factor β (which inhibits differentiation) allows calpastatin synthesis and prevents the diminution in calpastatin mRNA. The overall results suggest that at the onset of myoblast differentiation, calpastatin is regulated mainly at the level of translation and that an inhibition of calpastatin synthesis leads to the decrease in its mRNA stability. The existing calpastatin then diminishes, resulting in decreased calpastatin activity in the fusing myoblasts, allowing calpain activation and protein degradation required for fusion.

scholarly journals Granulosal and thecal expression of bone morphogenetic protein- and activin-binding protein mRNA transcripts during bovine follicle development and factors modulating their expression in vitro

Reproduction ◽  
2011 ◽  
Vol 142 (4) ◽  
pp. 581-591 ◽  
Author(s):  
Claire Glister ◽  
Leanne Satchell ◽  
Phil G Knight
Keyword(s):  
Bone Morphogenetic Proteins ◽  
Transforming Growth Factor ◽  
Ex Vivo ◽  
Mrna Level ◽  
Transcript Abundance ◽  
Transforming Growth Factor Β ◽  
Follicle Development ◽  
Mrna Levels ◽  
Theca Interna

Evidence supports local roles for transforming growth factor β superfamily members including activins and bone morphogenetic proteins (BMP) in follicle development. Access of these ligands to signalling receptors is likely modulated by extracellular binding proteins (BP). In this study, we comparedex vivoexpression of four BPs (chordin, gremlin, noggin and follistatin) in granulosal (GC) and theca interna (TC) compartments of developing bovine antral follicles (1–18 mm). Effects of FSH and IGF on BMP and BP expression by cultured GC, and effects of LH and BMPs on BP expression by cultured TC were also examined. Follicular expression of all four BP transcripts was higher in GC than TC compartments (P<0.001) a finding confirmed by immunohistochemistry. Follicle category affected (P<0.01) gremlin and follistatin mRNA abundance, with a significant cell-type×follicle category interaction for chordin, follistatin and noggin. Noggin transcript abundance was lower (P<0.05) in GC of large ‘E-active’ than ‘E-inactive’ follicles while follistatin mRNA level was higher (P<0.01). FSH enhanced CYP19, FSHR, INHBA and follistatin by GC without affecting BMP or BMP–BP expression. IGF increased CYP19 and follistatin, reduced BMP4, noggin and gremlin but did not affect chordin orFSHRmRNA levels. LH increased TC androgen secretion but had no effect on BMP or BP expression. BMPs uniformly suppressed TC androgen production whilst increasing chordin, noggin and gremlin mRNA levels up to 20-fold (P<0.01). These findings support the hypothesis that extracellular BP, mostly from GC, contribute to the regulation of intrafollicular BMP/activin signalling. Enhancement of thecal BP expression by BMP implies an autoregulatory feedback role to prevent excessive signalling.

Co-activation of synovial fibroblasts by laminin-111 and transforming growth factor-β induces expression of matrix metalloproteinases 3 and 10 independently of nuclear factor-κB

2007 ◽  
Vol 67 (4) ◽  
pp. 559-562 ◽  
Author(s):  
K Warstat ◽  
T Pap ◽  
G Klein ◽  
S Gay ◽  
W K Aicher
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Synovial Fibroblasts ◽  
Transforming Growth Factor Β ◽  
Nuclear Factor Κb ◽  
Mrna Levels ◽  
Nuclear Factor ◽  
The Matrix ◽  
Specific Tissue ◽  
Significant Expression

We showed previously that the attachment of synovial fibroblasts to laminin (LM)-111 in the presence of transforming growth factor-β induces significant expression of the matrix metalloproteinase (MMP)-3. Here we go on to investigate the regulation of additional MMPs and their specific tissue inhibitors of matrix proteases (TIMPs). Changes in steady-state mRNA levels encoding TIMPs and MMPs were investigated by quantitative reverse transcription–polymerase chain reaction. Production of MMPs was monitored by a multiplexed immunoarray. Signal transduction pathways were studied by immunoblotting. Attachment of synovial fibroblasts to LM-111 in the presence of transforming growth factor-β induced significant increases in MMP-3 mRNA (12.35-fold, p<0.001) and protein (mean 62 ng/ml, sixfold, p<0.008) and in expression of MMP-10 mRNA (11.68-fold, p<0.05) and protein (54 ng/ml, 20-fold, p⩾0.02). All other TIMPs and MMPs investigated failed to show this LM-111-facilitated transforming growth factor-β response. No phosphorylation of nuclear factor-κB was observed. We conclude that co-stimulation of synovial fibroblasts by LM-111 together with transforming growth factor-β suffices to induce significant expression of MMP-3 and MMP-10 by synovial fibroblasts and that this induction is independent of nuclear factor-κB phosphorylation.

scholarly journals Inhibition of Smurf2 translation by miR-322/503 modulates TGF-β/Smad2 signaling and intestinal epithelial homeostasis

2014 ◽  
Vol 25 (8) ◽  
pp. 1234-1243 ◽  
Author(s):  
Shan Cao ◽  
Lan Xiao ◽  
Jaladanki N. Rao ◽  
Tongtong Zou ◽  
Lan Liu ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Cellular Responses ◽  
Mrna Levels ◽  
Intestinal Epithelial ◽  
Regulatory Factor ◽  
Epithelial Homeostasis ◽  
Repressive Effect ◽  
Single Binding Site

Smad ubiquitin regulatory factor 2 (Smurf2) is an E3 ubiquitin ligase that regulates transforming growth factor β (TGF-β)/Smad signaling and is implicated in a wide variety of cellular responses, but the exact mechanisms that control Smurf2 abundance are largely unknown. Here we identify microRNA-322 (miR-322) and miR-503 as novel factors that regulate Smurf2 expression posttranscriptionally. Both miR-322 and miR-503 interact with Smurf2 mRNA via its 3′-untranslated region (UTR) and repress Smurf2 translation but do not affect total Smurf2 mRNA levels. Studies using heterologous reporter constructs reveal a greater repressive effect of miR-322/503 through a single binding site in the Smurf2 3′-UTR, whereas point mutation of this site prevents miR-322/503–induced repression of Smurf2 translation. Increased levels of endogenous Smurf2 via antagonism of miR-322/503 inhibits TGF-β–induced Smad2 activation by increasing degradation of phosphorylated Smad2. Furthermore, the increase in Smurf2 in intestinal epithelial cells (IECs) expressing lower levels of miR-322/503 is associated with increased resistance to apoptosis, which is abolished by Smurf2 silencing. These findings indicate that miR-322/503 represses Smurf2 translation, in turn affecting intestinal epithelial homeostasis by altering TGF-β/Smad2 signaling and IEC apoptosis.

scholarly journals The expression of transforming growth factor β in pregnant rat myometrium is hormone and stretch dependent

Reproduction ◽  
2007 ◽  
Vol 134 (3) ◽  
pp. 503-511 ◽  
Author(s):  
Oksana Shynlova ◽  
Prudence Tsui ◽  
Anna Dorogin ◽  
B Lowell Langille ◽  
Stephen J Lye
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Late Gestation ◽  
Quiescent State ◽  
Mrna Levels ◽  
Pregnant Rats ◽  
Pregnant Rat ◽  
Threefold Increase ◽  
Antagonist Ru486

From a quiescent state in early pregnancy to a highly contractile state in labor, the myometrium displays tremendous growth and remodeling. We hypothesize that the transforming growth factor β (TGFβ) system is involved in the differentiation of pregnant myometrium throughout gestation and labor. Furthermore, we propose that during pregnancy the mechanical and hormonal stimuli play a role in regulating myometrial TGFβs. The expression of TGFβ1-3 mRNAs and proteins was examined by real-time PCR, Western immunoblot, and localized with immunohistochemistry in the rat uterus throughout pregnancy and labor. Tgfβ1-3 genes were expressed differentially in pregnant myometrium. Tgfβ2 gene was not affected by pregnancy, whereas the Tgfβ1 gene showed a threefold increase during the second half of gestation. In contrast, we observed a dramatic bimodal change in Tgfβ3 gene expression throughout pregnancy. Tgfβ3 mRNA levels first transiently increased at mid-gestation (11-fold on day 14) and later at term (45-fold at labor, day 23). Protein expression levels paralleled the changes in mRNA. Treatment of pregnant rats with the progesterone (P4) receptor antagonist RU486 induced premature labor on day 19 and increased Tgfβ3 mRNA, whereas artificial maintenance of elevated P4 levels at late gestation (days 20–23) caused a significant decrease in the expression of Tgfβ3 gene. In addition, Tgfβ3 was up-regulated specifically in the gravid horn of unilaterally pregnant rats subjected to a passive biological stretch imposed by the growing fetuses, but not in the empty horn. Collectively, these data indicate that the TGFβ family contributes in the regulation of myometrial activation at term integrating mechanical and endocrine signals for successful labor contraction.

Abstract 297: Cardiac Transforming-growth-factor β-stimulated Clone 22 Gene Expression By Hypertrophic Stimuli

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Annina Kelloniemi ◽  
Jani Aro ◽  
Elina Koivisto ◽  
Heikki Ruskoaho ◽  
Jaana Rysä
Keyword(s):  
Growth Factor ◽  
Gene Transfer ◽  
Transforming Growth Factor ◽  
Pressure Overload ◽  
Transforming Growth Factor Β ◽  
Left Ventricular ◽  
Neonatal Rat ◽  
Mrna Levels ◽  
Ang Ii ◽  
Sd Rats

Objectives: Transforming-growth-factor β-stimulated clone 22 (TSC-22) is a leucine zipper protein expressed in many tissues and possessing various transcription-modulating activities. However, its function in the heart remains largely unknown. The aim of the present study was to characterize the cardiac TSC-22 expression. Methods: Acute pressure overload was accomplished in conscious Sprague-Dawley (SD) rats by intravenous infusion of arginine 8 -vasopressin (AVP, 0.05 μg/kg/min) for 4 hours and subcutaneous infusion of angiotensin II (Ang II, 33 μg/kg/h) with and without Ang II receptor type 1 blocker losartan (400 μg/kg/h) by using osmotic minipumps for 2 weeks. Adenovirus-mediated intramyocardial gene transfer of TSC-22 was performed into left ventricle (LV) of SD rats. Experimental myocardial infarction (MI) was produced by ligation of the left anterior descending coronary artery. Cultured neonatal rat ventricular myocytes (NRVM) were treated with endothelin-1 (ET-1, 100 nM). Results: A significant 1.6-fold increase ( P <0.05) in LV TSC-22 mRNA levels was noted already after 1 hour AVP infusion. Moreover, Ang II infusion markedly upregulated TSC-22 expression, LV mRNA levels being highest at 6 hours (11-fold, P <0.001). Simultaneous infusion of losartan completely abolished Ang II-induced increase in TSC-22 mRNA levels. Adenovirus-mediated gene transfer of TSC-22 into LV resulted a 1.9-fold ( P <0.001) increase in TSC-22 mRNA levels, accompanied by upregulated BNP mRNA levels (1.4-fold, P <0.01). In response to experimental MI, TSC-22 mRNA levels were elevated 4.1-fold ( P <0.001) at 1 day and 1.9-fold ( P <0.05) at 4 weeks. In cultured NRVM, ET-1 treatment increased TSC-22 mRNA levels from 1 h to 24 h, the greatest increase being observed at 12 h (2.7-fold, P <0.001). TSC-22 protein levels were upregulated from 4 h to 24 h with the highest increase at 24 h (4.7-fold, P <0.01). Conclusion: These results indicate that TSC-22 expression is rapidly activated in response to pressure overload, MI and in ET-1 treated cultured NRVM. Moreover, adenovirus-mediated overexpression of TSC-22 mRNA was associated with elevated left ventricular BNP mRNA levels.

scholarly journals Transforming Growth Factor-βs Inhibit Somatostatin Messenger Ribonucleic Acid Levels and Somatostatin Secretion in Hypothalamic Cells in Culture*

Endocrinology ◽  
1997 ◽  
Vol 138 (10) ◽  
pp. 4401-4409 ◽  
Author(s):  
M. Quintela ◽  
R. M. SeñarÍs ◽  
C. Diéguez
Keyword(s):  
Growth Factor ◽  
Messenger Rna ◽  
Transforming Growth Factor ◽  
Inhibitory Effect ◽  
Mrna Levels ◽  
Messenger Ribonucleic Acid ◽  
Cells In Culture ◽  
Somatostatin Secretion ◽  
Hypothalamic Cells ◽  
The Stability

Abstract Treatment of hypothalamic cells in monolayer culture with transforming growth factor-β1 (TGFβ1) significantly reduced both basal and cAMP-induced somatostatin messenger RNA (mRNA) levels and somatostatin secretion. This inhibitory effect was dose- and time-dependent and not mediated by glial cells, as it was also observed in glial-free hypothalamic cell cultures treated with cytosine arabinonucleoside. TGFβ2 and -β3 mimicked the actions of TGFβ1, which indicated that the three isoforms of the TGFβ family expressed in the central nervous system displayed similar effects on the somatostatinergic neurons. The blockade of synthesis of proteins with either cycloheximide or puromycin for 24 h prevented the inhibitory effect of TGFβ1 on somatostatin mRNA. This implied that the reduction of this mRNA by TGFβ1 required de novo protein synthesis. We next studied whether TGFβ1 acted at the transcriptional or posttranscriptional level by altering the stability of somatostatin mRNA. Examination of the rate of disappearance of somatostatin mRNA by Northern blot, after inhibition of mRNA transcription with either actinomycin D (AcD) or 5,6-dichloro-1β-ribofuranosyl benzimidazole revealed that TGFβ1 did reduce the stability of somatostatin mRNA. This effect was observed when we pretreated the cultures with TGFβ1 4 h before the addition of AcD, but not when we administered TGFβ1 simultaneously with AcD or 5,6-dichloro-1β-ribofuranosyl benzimidazole. Altogether these results demonstrated that the treatment of hypothalamic cells in culture with TGFβ1, TGFβ2, or TGFβ3 resulted in a decrease in somatostatin mRNA levels and somatostatin secretion. TGFβ1 reduced the steady state levels of somatostatin mRNA by inducing the synthesis of a protein (s), that appears to accelerate the degradation of the mRNA of somatostatin. Whether TGFβ1 has additional effects on the transcription of the somatostatin gene will require further study.

scholarly journals Pioglitazone attenuates hepatic inflammation and fibrosis in phosphatidylethanolamine N-methyltransferase-deficient mice

2016 ◽  
Vol 310 (7) ◽  
pp. G526-G538 ◽  
Author(s):  
Jelske N. van der Veen ◽  
Susanne Lingrell ◽  
Xia Gao ◽  
Ariel D. Quiroga ◽  
Abhijit Takawale ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Transforming Growth Factor ◽  
Body Weight Gain ◽  
Smooth Muscle Actin ◽  
Transforming Growth Factor Β ◽  
Liver Weight ◽  
Mrna Levels ◽  
Nonalcoholic Fatty Liver ◽  
Diet Induced Obesity ◽  
Important Enzyme

Phosphatidylethanolamine N-methyltransferase (PEMT) is an important enzyme in hepatic phosphatidylcholine (PC) biosynthesis. Pemt−/− mice are protected against high-fat diet (HFD)-induced obesity and insulin resistance; however, these mice develop nonalcoholic fatty liver disease (NAFLD). We hypothesized that peroxisomal proliferator-activated receptor-γ (PPARγ) activation by pioglitazone might stimulate adipocyte proliferation, thereby directing lipids from the liver toward white adipose tissue. Pioglitazone might also act directly on PPARγ in the liver to improve NAFLD. Pemt+/+ and Pemt−/− mice were fed a HFD with or without pioglitazone (20 mg·kg−1·day−1) for 10 wk. Pemt−/− mice were protected from HFD-induced obesity but developed NAFLD. Treatment with pioglitazone caused an increase in body weight gain in Pemt−/− mice that was mainly due to increased adiposity. Moreover, pioglitazone improved NAFLD in Pemt−/− mice, as indicated by a 35% reduction in liver weight and a 57% decrease in plasma alanine transaminase levels. Livers from HFD-fed Pemt−/− mice were steatotic, inflamed, and fibrotic. Hepatic steatosis was still evident in pioglitazone-treated Pemt−/− mice; however, treatment with pioglitazone reduced hepatic fibrosis, as evidenced by reduced Sirius red staining and lowered mRNA levels of collagen type Iα1 ( Col1a1), tissue inhibitor of metalloproteinases 1 ( Timp1), α-smooth muscle actin ( Acta2), and transforming growth factor-β ( Tgf-β). Similarly, oxidative stress and inflammation were reduced in livers from Pemt−/− mice upon treatment with pioglitazone. Together, these data show that activation of PPARγ in HFD-fed Pemt −/− mice improved liver function, while these mice were still protected against diet-induced obesity and insulin resistance.

The anemia of the newborn induces erythropoietin expression in the developing mouse retina

2010 ◽  
Vol 299 (1) ◽  
pp. R111-R118 ◽  
Author(s):  
N. Scheerer ◽  
N. Dünker ◽  
S. Imagawa ◽  
M. Yamamoto ◽  
N. Suzuki ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
Fluorescent Protein ◽  
Ganglion Cells ◽  
Retinal Development ◽  
Transforming Growth Factor Β ◽  
Neuroprotective Activity ◽  
Mouse Retina ◽  
Mrna Levels ◽  
Neuroprotective Effects

The hematopoietic hormone erythropoietin (Epo), regularly produced by the kidneys and the liver, is also expressed in neuronal tissue, where it has been found to mediate paracrine neuroprotective effects. In most studies exploring the rescue effects of Epo, apoptosis was exogenously induced by different cell death stimuli. Herein, we set out to study the expression and function of Epo in physiologically occurring apoptosis in a model of retinal development. We made use of an organotypic retinal wholemount culture system that resembles the physiological in vivo situation with cell connections still retained. Epo mRNA expression in the retina, liver, and kidney showed a significant increase during early development, coinciding with the anemia of the newborn. In the retina of Epo-green fluorescent protein transgenic mice, Epo-expressing cells were identified and found to be distributed in the retinal ganglion cell layer. Treatment of retinal wholemount cultures with recombinant Epo resulted in a significant decrease of apoptotic ganglion cells as well as photoreceptor cells throughout retinal development. Moreover, transforming growth factor-β-induced apoptosis was completely antagonized by Epo when both factors were simultaneously applied. Investigations on the signaling pathway revealed a decrease in Bax mRNA levels in Epo-treated retinal cells. We conclude that Epo exerts wide and prolonged neuroprotective activity in physiologically occurring apoptosis and thus contributes to proper retinal development.

Acute regulation of activin A and its binding protein, follistatin, in serum and tissues following lipopolysaccharide treatment of adult male mice

2012 ◽  
Vol 303 (6) ◽  
pp. R665-R675 ◽  
Author(s):  
Hui Wu ◽  
Yi Chen ◽  
Wendy R. Winnall ◽  
David J. Phillips ◽  
Mark P. Hedger
Keyword(s):  
Bone Marrow ◽  
Adult Male ◽  
Transforming Growth Factor ◽  
Binding Protein ◽  
Protein Translation ◽  
Transforming Growth Factor Β ◽  
Activin A ◽  
Male Mice ◽  
Mrna Transcription ◽  
Bone Marrow Derived Cells

Activin A, a member of the transforming growth factor-β family, increases in the circulation within 1 h after administration of bacterial LPS. To clarify the origins of this rapid increase, the distribution of activin A and its binding protein, follistatin, and their production following LPS treatment, were assessed in adult male mice. In untreated mice, activin A was detectable in all 23 tissues examined, with highest mRNA expression (as measured by quantitative RT-PCR) was found in the liver, and the largest concentration of activin A protein (by ELISA) was found in the bone marrow. Likewise, follistatin mRNA and protein were present in all tissues, with highest expression in the vas deferens. Activin A and follistatin mRNA did not increase significantly in any tissue within the first hour after LPS, but activin A protein decreased by 35% in the bone marrow and increased 5-fold in the lung. No significant changes were observed in any other tissue. Activin A reached a peak in the circulation 1 h following LPS, and then declined. Cycloheximide, an inhibitor of protein translation, reduced this increase of activin A by more than 50%. Actinomycin D, an inhibitor of mRNA transcription, had no effect. Circulating follistatin did not increase until 4 h after LPS and was not affected by either inhibitor. These data indicate that the rapid increase in circulating activin A during LPS-induced inflammation is regulated at the posttranscriptional level, apparently from newly translated and stored protein, and implicate bone marrow-derived cells, and, in particular, neutrophils, as a significant source of this preformed activin A.

TGF-β and CTGF have overlapping and distinct fibrogenic effects on human renal cells

2002 ◽  
Vol 283 (4) ◽  
pp. F707-F716 ◽  
Author(s):  
Elizabeth Gore-Hyer ◽  
Daniel Shegogue ◽  
Malgorzata Markiewicz ◽  
Shianlen Lo ◽  
Debra Hazen-Martin ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Growth Factors ◽  
Growth Factor ◽  
Renal Fibrosis ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Tissue Growth ◽  
Mrna Levels ◽  
Potent Inducer ◽  
Collagen Promoter

Transforming growth factor-β (TGF-β) and connective tissue growth factor (CTGF) are ubiquitously expressed in various forms of tissue fibrosis, including fibrotic diseases of the kidney. To clarify the common and divergent roles of these growth factors in the cells responsible for pathological extracellular matrix (ECM) deposition in renal fibrosis, the effects of TGF-β and CTGF on ECM expression in primary human mesangial (HMCs) and human proximal tubule epithelial cells (HTECs) were studied. Both TGF-β and CTGF significantly induced collagen protein expression with similar potency in HMCs. Additionally, α2(I)-collagen promoter activity and mRNA levels were similarly induced by TGF-β and CTGF in HMCs. However, only TGF-β stimulated collagenous protein synthesis in HTECs. HTEC expression of tenascin-C (TN-C) was increased by TGF-β and CTGF, although TGF-β was the more potent inducer. Thus both growth factors elicit similar profibrogenic effects on ECM production in HMCs, while promoting divergent effects in HTECs. CTGF induction of TN-C, a marker of epithelial-mesenchymal transdifferentiation (EMT), with no significant induction of collagenous protein synthesis in HTECs, may suggest a more predominant role for CTGF in EMT rather than induction of excessive collagen deposition by HTECs during renal fibrosis.

Sign in / Sign up

Export Citation Format

Share Document