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

Barnoy, Sivia; SUPINO-ROSIN, Lia; Kosower, Nechama S.

doi:10.1042/0264-6021:3510413

Cited by 27 publications

(18 citation statements)

References 21 publications

(34 reference statements)

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“…For example, calciumdependent muscle proteolysis in tumor-bearing rats was found to reflect reduced calpastatin activity in a recent study by Costelli et al (11). In other studies, calpain-dependent protein degradation in differentiating myoblasts reflected reduced calpastatin expression and increased calpain-to-calpastatin ratio (2).…”

Section: Discussionmentioning

confidence: 97%

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

Wei¹,

Fareed²,

Evenson³

et al. 2005

American Journal of Physiology-Regulatory, Integrative and Comp

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, and Per-Olof Hasselgren. Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3. Am J Physiol Regul Integr Comp Physiol 288: R580 -R590, 2005. First published November 24, 2004 doi:10.1152/ajpregu.00341.2004.-We examined the influence of sepsis on the expression and activity of the calpain and caspase systems in skeletal muscle. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham operated. Calpain activity was determined by measuring the calcium-dependent hydrolysis of casein and by casein zymography. The activity of the endogenous calpain inhibitor calpastatin was measured by determining the inhibitory effect on calpain activity in muscle extracts. Protein levels of -and m-calpain and calpastatin were determined by Western blotting, and calpastatin mRNA was measured by real-time PCR. Caspase-3 activity was determined by measuring the hydrolysis of the fluorogenic caspase-3 substrate Ac-DEVD-AMC and by determining protein and mRNA expression for caspase-3 by Western blotting and real-time PCR, respectively. In addition, the role of calpains and caspase-3 in sepsis-induced muscle protein breakdown was determined by measuring protein breakdown rates in the presence of specific inhibitors. Sepsis resulted in increased muscle calpain activity caused by reduced calpastatin activity. In contrast, caspase-3 activity, mRNA levels, and activated caspase-3 29-kDa fragment were not altered in muscle from septic rats. Sepsis-induced muscle proteolysis was blocked by the calpain inhibitor calpeptin but was not influenced by the caspase-3 inhibitor Ac-DEVD-CHO. The results suggest that sepsis-induced muscle wasting is associated with increased calpain activity, secondary to reduced calpastatin activity, and that caspase-3 activity is not involved in the catabolic response to sepsis.

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Section: Discussionmentioning

confidence: 97%

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

Wei¹,

Fareed²,

Evenson³

et al. 2005

American Journal of Physiology-Regulatory, Integrative and Comp

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“…Under normal conditions the major calpain isozymes and the endogenous calpain inhibitor calpastatin have been reported to exist as relatively stable, long-lived proteins (4,64). However, in response to particular stimuli, such as ischemic or reperfusion injury in vivo or elevation of intracellular calcium levels, in vitro calpastatin acts as a substrate for calpain-mediated degradation (9,40,44,46,56).…”

Section: Discussionmentioning

confidence: 99%

v-Src-Induced Modulation of the Calpain-Calpastatin Proteolytic System Regulates Transformation

Carragher

Westhoff

Riley

et al. 2002

Molecular and Cellular Biology

105

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v-Src-induced oncogenic transformation is characterized by alterations in cell morphology, adhesion, motility, survival, and proliferation. To further elucidate some of the signaling pathways downstream of v-Src that are responsible for the transformed cell phenotype, we have investigated the role that the calpain-calpastatin proteolytic system plays during oncogenic transformation induced by v-Src. We recently reported that v-Srcinduced transformation of chicken embryo fibroblasts is accompanied by calpain-mediated proteolytic cleavage of the focal adhesion kinase (FAK) and disassembly of the focal adhesion complex. In this study we have characterized a positive feedback loop whereby activation of v-Src increases protein synthesis of calpain II, resulting in degradation of its endogenous inhibitor calpastatin. Reconstitution of calpastatin levels by overexpression of exogenous calpastatin suppresses proteolytic cleavage of FAK, morphological transformation, and anchorage-independent growth. Furthermore, calpastatin overexpression represses progression of v-Src-transformed cells through the G 1 stage of the cell cycle, which correlates with decreased pRb phosphorylation and decreased levels of cyclins A and D and cyclin-dependent kinase 2. Calpain 4 knockout fibroblasts also exhibit impaired v-Src-induced morphological transformation and anchorage-independent growth. Thus, modulation of the calpain-calpastatin proteolytic system plays an important role in focal adhesion disassembly, morphological transformation, and cell cycle progression during v-Src-induced cell transformation.Oncogenic transformation of cells by v-Src is associated with deregulated growth control, cytoskeketal disassembly, and loss of integrin-linked focal adhesion structures (17,20,27,31). Such alterations contribute to the highly mitogenic and motile phenotype that characterizes v-Src transformation. The precise mechanisms by which v-Src promotes cell transformation remain poorly understood. Previous studies, however, indicate that v-Src-induced morphological transformation occurs by mechanisms independent of gene expression (8,22), implicating Src kinase activity or other posttranscriptional mechanisms as key mediators of v-Src-induced transformation. Calpainmediated proteolysis represents a major pathway of posttranslational modification of cellular proteins and has been implicated in diverse cellular processes ranging from apoptosis to cell migration and cell cycle progression (12,25,43,45,49,57). We have previously demonstrated that calpain-mediated proteolytic cleavage of the focal adhesion kinase (FAK) and focal adhesion disassembly accompany v-Src-induced morphological transformation. Calpain-mediated disassembly of focal adhesions results in a reduction in the strength of adhesion that transformed cells have to their culture substrate, thereby promoting cell motility (12).The calpains represent a highly conserved family of nonlysosomal calcium-dependent cysteine proteases comprising two ubiquitously expressed isoforms, -calpain (cal...

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“…Calpastatin is a specific endogenous inhibitor of the protease calpain and acts by binding to calpain (Sorimachi et al, 1997). The balance between calpain proteolysis and calpastatin inhibition regulates stability of the cytoskeleton in a variety of circumstances (Barnoy et al, 1998;Barnoy et al, 2000;Croce et al, 1999;Dwyer-Nield et al, 1996;Potter et al, 1998). Calpastatin and the various calpain isoforms are ubiquitous proteins present in a wide range of cell types.…”

Section: Discussionmentioning

confidence: 99%

Foxj1 regulates basal body anchoring to the cytoskeleton of ciliated pulmonary epithelial cells

Gomperts

Gong-Cooper

Hackett

2004

Journal of Cell Science

123

102

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The forkhead box transcription factor Foxj1 is required for cilia formation and left-right axis determination. To define the role of Foxj1 in ciliogenesis, microarray analysis was performed to identify differentially expressed genes in the pulmonary epithelium of foxj1 +/+ and foxj1 -/-mice. In the absence of Foxj1, the expression of calpastatin, an inhibitor of the protease calpain, decreased. RNase protection confirmed the decrease in calpastatin expression and decreased calpastatin was detected in the proximal pulmonary epithelium of foxj1 -/-mice by immunohistochemistry. No change was detected in the expression of calpain 2 in the pulmonary epithelium by western blot or immunohistochemistry. By western blot and immunofluorescence, ezrin, a substrate for calpain, was also found to decrease in the pulmonary epithelium of foxj1 -/-mice. No change in ezrin gene expression was found by RT-PCR. A decrease in ezrin binding phosphoprotein-50 (EBP-50) was also detected by immunofluorescence in the foxj1 -/-mouse pulmonary epithelium. Immunoelectron microscopy demonstrated ezrin associated with the basal bodies of cilia in the pulmonary epithelium. Treatment of tracheal explants from foxj1 -/-mice with a calpain inhibitor resulted in a partial reappearance of cilia observed in these mice. Additionally, following treatment of foxj1 -/-tracheal explants with calpain inhibitor, basal bodies were observed in an apical location along with relocalization of ezrin and EBP-50. Regulation of calpain activity by calpastatin thus provides a mechanism for regulating the anchoring of basal bodies to the apical cytoskeleton in ciliated cells. In the absence of Foxj1, decreased calpastatin expression with decreased ezrin and EBP-50 results in an inability of basal bodies to anchor to the apical cytoskeleton and subsequent failure of axonemal formation.

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

Cited by 27 publications

References 21 publications

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

v-Src-Induced Modulation of the Calpain-Calpastatin Proteolytic System Regulates Transformation

Foxj1 regulates basal body anchoring to the cytoskeleton of ciliated pulmonary epithelial cells

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