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CIP4 is required for the hypertrophic growth of neonatal cardiac myocytes.

Rusconi F, Thakur H, Li J, Kapiloff MS - J. Biomed. Sci. (2013)

Bottom Line: CIP4 is a scaffold protein that regulates membrane deformation and tubulation, organization of the actin cytoskeleton, endocytosis of growth factor receptors, and vesicle trafficking.Although expressed in the heart, CIP4 has not been studied with regards to its potential function in cardiac myocytes.These results imply that CIP4 plays a significant role in the intracellular hypertrophic signal transduction network that controls the growth of cardiac myocytes in heart disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interdisciplinary Stem Cell Institute, Department of Pediatrics, Leonard M. Miller School of Medicine, University of Miami, Miami, FL 33101, USA.

ABSTRACT

Background: CIP4 is a scaffold protein that regulates membrane deformation and tubulation, organization of the actin cytoskeleton, endocytosis of growth factor receptors, and vesicle trafficking. Although expressed in the heart, CIP4 has not been studied with regards to its potential function in cardiac myocytes.

Results: We now show using RNA interference that CIP4 expression in neonatal rat ventricular myocytes is required for the induction of non-mitotic, hypertrophic growth by the α-adrenergic agonist phenylephrine, the IL-6 cytokine leukemia inhibitor factor, and fetal bovine serum, as assayed using morphometry, immunocytochemistry for the hypertrophic marker atrial natriuretic factor and [3H]leucine incorporation for de novo protein synthesis. This requirement was consistent with the induction of CIP4 expression by hypertrophic stimulation. The inhibition of myocyte hypertrophy by CIP4 small interfering oligonucleotides (siRNA) was rescued by expression of a recombinant CIP4 protein, but not by a mutant lacking the N-terminal FCH domain responsible for CIP4 intracellular localization.

Conclusions: These results imply that CIP4 plays a significant role in the intracellular hypertrophic signal transduction network that controls the growth of cardiac myocytes in heart disease.

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CIP4 structure and expression. A. Primary structure of CIP4 showing domains and binding sites for known protein partners. “h” indicates the 56 amino acid residue insertion present in CIP4h. B. Neonatal rat ventricular myocytes were transfected with control or CIP4 siRNA and then stimulated with 10 μM PE, 1000 U/mL LIF, or 10% FBS for two days as indicated. CIP4 proteins in whole cell lysates were detected using a mouse anti-CIP4 antibody. n = 4–5. †p-values vs. no drug control; *p-values vs. control siRNA-transfected myocytes treated with the same agonist. C. Myocytes were transfected with control or CIP4 siRNA and cultured in minimal media +/− 4% horse serum for two days before TUNEL staining. % TUNEL-positive nuclei are indicated. **p < 0.005 for CIP4 siRNA vs. control siRNA. n = 3–5.
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Figure 1: CIP4 structure and expression. A. Primary structure of CIP4 showing domains and binding sites for known protein partners. “h” indicates the 56 amino acid residue insertion present in CIP4h. B. Neonatal rat ventricular myocytes were transfected with control or CIP4 siRNA and then stimulated with 10 μM PE, 1000 U/mL LIF, or 10% FBS for two days as indicated. CIP4 proteins in whole cell lysates were detected using a mouse anti-CIP4 antibody. n = 4–5. †p-values vs. no drug control; *p-values vs. control siRNA-transfected myocytes treated with the same agonist. C. Myocytes were transfected with control or CIP4 siRNA and cultured in minimal media +/− 4% horse serum for two days before TUNEL staining. % TUNEL-positive nuclei are indicated. **p < 0.005 for CIP4 siRNA vs. control siRNA. n = 3–5.

Mentions: CIP4 is a modular scaffold protein involved in the regulation of cellular morphology that can serve as an effector for the Rho family small GTPases Cdc42, TC10, and TCL [3]. CIP4 contains a N-terminal F-Bar (Fes-CIP4 homology [FCH] – Bin/Amphyphysin/Rvs) domain that binds both cytoskeletal proteins and negatively-charged membrane phospholipids, a HR1 domain that binds active, GTP-bound Rho family members, and a C-terminal SH3 (SRC Homology 3) domain that binds a variety of proteins involved in the regulation of the actin cytoskeleton and small GTPase signaling (Figure 1A). As a result, CIP4 has roles in the regulation of membrane deformation and tubulation, dynamic remodeling of the actin cytoskeleton, endocytosis, and vesicle trafficking [4,5]. CIP4 also regulates filopodial and lamellipodial protrusion, affecting the invasiveness and metastasis of cancer cells and neurite extension in neurons [6,7]. Recently, two groups described CIP4 knock-out mice that were normal in appearance and fertile. One group showed that CIP4- embryonic fibroblasts were defective in endocytosis [4]. This was reflected by increased GLUT4 (glucose transporter 4) levels in skeletal muscle membranes and lower post-prandial glucose levels in vivo. The other group showed that despite normal development of the immune system, the CIP4- mice had poor T-cell function [5]. These defects were related to impaired T cell migration and adhesion, presumably reflecting the importance of CIP4 to regulation of the actin cytoskeleton.


CIP4 is required for the hypertrophic growth of neonatal cardiac myocytes.

Rusconi F, Thakur H, Li J, Kapiloff MS - J. Biomed. Sci. (2013)

CIP4 structure and expression. A. Primary structure of CIP4 showing domains and binding sites for known protein partners. “h” indicates the 56 amino acid residue insertion present in CIP4h. B. Neonatal rat ventricular myocytes were transfected with control or CIP4 siRNA and then stimulated with 10 μM PE, 1000 U/mL LIF, or 10% FBS for two days as indicated. CIP4 proteins in whole cell lysates were detected using a mouse anti-CIP4 antibody. n = 4–5. †p-values vs. no drug control; *p-values vs. control siRNA-transfected myocytes treated with the same agonist. C. Myocytes were transfected with control or CIP4 siRNA and cultured in minimal media +/− 4% horse serum for two days before TUNEL staining. % TUNEL-positive nuclei are indicated. **p < 0.005 for CIP4 siRNA vs. control siRNA. n = 3–5.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750294&req=5

Figure 1: CIP4 structure and expression. A. Primary structure of CIP4 showing domains and binding sites for known protein partners. “h” indicates the 56 amino acid residue insertion present in CIP4h. B. Neonatal rat ventricular myocytes were transfected with control or CIP4 siRNA and then stimulated with 10 μM PE, 1000 U/mL LIF, or 10% FBS for two days as indicated. CIP4 proteins in whole cell lysates were detected using a mouse anti-CIP4 antibody. n = 4–5. †p-values vs. no drug control; *p-values vs. control siRNA-transfected myocytes treated with the same agonist. C. Myocytes were transfected with control or CIP4 siRNA and cultured in minimal media +/− 4% horse serum for two days before TUNEL staining. % TUNEL-positive nuclei are indicated. **p < 0.005 for CIP4 siRNA vs. control siRNA. n = 3–5.
Mentions: CIP4 is a modular scaffold protein involved in the regulation of cellular morphology that can serve as an effector for the Rho family small GTPases Cdc42, TC10, and TCL [3]. CIP4 contains a N-terminal F-Bar (Fes-CIP4 homology [FCH] – Bin/Amphyphysin/Rvs) domain that binds both cytoskeletal proteins and negatively-charged membrane phospholipids, a HR1 domain that binds active, GTP-bound Rho family members, and a C-terminal SH3 (SRC Homology 3) domain that binds a variety of proteins involved in the regulation of the actin cytoskeleton and small GTPase signaling (Figure 1A). As a result, CIP4 has roles in the regulation of membrane deformation and tubulation, dynamic remodeling of the actin cytoskeleton, endocytosis, and vesicle trafficking [4,5]. CIP4 also regulates filopodial and lamellipodial protrusion, affecting the invasiveness and metastasis of cancer cells and neurite extension in neurons [6,7]. Recently, two groups described CIP4 knock-out mice that were normal in appearance and fertile. One group showed that CIP4- embryonic fibroblasts were defective in endocytosis [4]. This was reflected by increased GLUT4 (glucose transporter 4) levels in skeletal muscle membranes and lower post-prandial glucose levels in vivo. The other group showed that despite normal development of the immune system, the CIP4- mice had poor T-cell function [5]. These defects were related to impaired T cell migration and adhesion, presumably reflecting the importance of CIP4 to regulation of the actin cytoskeleton.

Bottom Line: CIP4 is a scaffold protein that regulates membrane deformation and tubulation, organization of the actin cytoskeleton, endocytosis of growth factor receptors, and vesicle trafficking.Although expressed in the heart, CIP4 has not been studied with regards to its potential function in cardiac myocytes.These results imply that CIP4 plays a significant role in the intracellular hypertrophic signal transduction network that controls the growth of cardiac myocytes in heart disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interdisciplinary Stem Cell Institute, Department of Pediatrics, Leonard M. Miller School of Medicine, University of Miami, Miami, FL 33101, USA.

ABSTRACT

Background: CIP4 is a scaffold protein that regulates membrane deformation and tubulation, organization of the actin cytoskeleton, endocytosis of growth factor receptors, and vesicle trafficking. Although expressed in the heart, CIP4 has not been studied with regards to its potential function in cardiac myocytes.

Results: We now show using RNA interference that CIP4 expression in neonatal rat ventricular myocytes is required for the induction of non-mitotic, hypertrophic growth by the α-adrenergic agonist phenylephrine, the IL-6 cytokine leukemia inhibitor factor, and fetal bovine serum, as assayed using morphometry, immunocytochemistry for the hypertrophic marker atrial natriuretic factor and [3H]leucine incorporation for de novo protein synthesis. This requirement was consistent with the induction of CIP4 expression by hypertrophic stimulation. The inhibition of myocyte hypertrophy by CIP4 small interfering oligonucleotides (siRNA) was rescued by expression of a recombinant CIP4 protein, but not by a mutant lacking the N-terminal FCH domain responsible for CIP4 intracellular localization.

Conclusions: These results imply that CIP4 plays a significant role in the intracellular hypertrophic signal transduction network that controls the growth of cardiac myocytes in heart disease.

Show MeSH
Related in: MedlinePlus