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Muscle ring finger protein-1 inhibits PKC{epsilon} activation and prevents cardiomyocyte hypertrophy.

Arya R, Kedar V, Hwang JR, McDonough H, Li HH, Taylor J, Patterson C - J. Cell Biol. (2004)

Bottom Line: Much effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy.MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1.MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size.

View Article: PubMed Central - PubMed

Affiliation: Carolina Cardiovascular Biology Center, University of North Carolina, Chapel Hill, NC 27599, USA.

ABSTRACT
Much effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy. In spite of this, we still know little about the mechanisms that inhibit hypertrophic growth. We define a novel anti-hypertrophic signaling pathway regulated by muscle ring finger protein-1 (MURF1) that inhibits the agonist-stimulated PKC-mediated signaling response in neonatal rat ventricular myocytes. MURF1 interacts with receptor for activated protein kinase C (RACK1) and colocalizes with RACK1 after activation with phenylephrine or PMA. Coincident with this agonist-stimulated interaction, MURF1 blocks PKCepsilon translocation to focal adhesions, which is a critical event in the hypertrophic signaling cascade. MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1. MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size. These findings establish that MURF1 is a key regulator of the PKC-dependent hypertrophic response and can blunt cardiomyocyte hypertrophy, which may have important implications in the pathophysiology of clinical cardiac hypertrophy.

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MURF1 inhibits PMA-induced translocation of PKCε from the perinuclear region to focal adhesion complexes in NRVM. NRVM were infected with Ad.GFP (A and B) or Ad.MURF1 (C and D) for 24 h in serum-free medium followed by induction with PMA (B and D) for 15 min. The cells were fixed and incubated with anti-vinculin (blue) and anti-PKCε (red) for 2 h. Arrows indicate focal adhesions.
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fig4: MURF1 inhibits PMA-induced translocation of PKCε from the perinuclear region to focal adhesion complexes in NRVM. NRVM were infected with Ad.GFP (A and B) or Ad.MURF1 (C and D) for 24 h in serum-free medium followed by induction with PMA (B and D) for 15 min. The cells were fixed and incubated with anti-vinculin (blue) and anti-PKCε (red) for 2 h. Arrows indicate focal adhesions.

Mentions: Because RACK1 is known to regulate targeting of PKC isoforms, we examined the translocation of PKC isoforms in NRVM in the presence of MURF1. In nonstimulated myocytes, PKCβII is associated with fibrillar structures and after activation it translocates to the nuclear and cell periphery, whereas before stimulation, PKCε is observed in the nucleus and perinucleus but translocates to cross-striated structures and focal adhesions in stimulated cells (Disatnik et al., 1994a; Ron et al., 1995; Johnson et al., 1996; Heidkamp et al., 2003). PKC activation induces the association and colocalization of RACK1 with PKCβII (Ron et al., 1999); MuRF1 modestly increased perinuclear localization of PKCβII under unstimulated conditions and did not inhibit PE- or PMA-dependent PKCβII translocation (Fig. 3). PKCβII moved from cytosol to perinuclear structures after activation with PE and PMA, and colocalized with RACK1 in control as well as Ad.MURF1-infected NRVM. Several recent reports have also shown colocalization of PKCε and RACK1 in response to stimulation with PE and PMA (Pass et al., 2001a,b; Besson et al., 2002). Using an anti-PKCε antibody, we observed the translocation of PKCε from perinuclear structures to focal adhesions upon activation with PE or PMA in NRVM infected with Ad.GFP, as indicated by costaining with the focal adhesion marker vinculin. Surprisingly, this translocation was inhibited after infection with Ad.MURF1 (Fig. 4). PKCε translocation occurred in 22 ± 6% of Ad.MURF1-infected cells treated with PMA compared with 82 ± 8% of control cells. After stimulation with PMA, PKCε was still observed in the perinuclear structures and nuclei in these cells. Similar patterns were observed after stimulation with PE (unpublished data). These data indicate that MURF1 specifically inhibits the movement of the PKCε isoform after agonist-induced stimulation.


Muscle ring finger protein-1 inhibits PKC{epsilon} activation and prevents cardiomyocyte hypertrophy.

Arya R, Kedar V, Hwang JR, McDonough H, Li HH, Taylor J, Patterson C - J. Cell Biol. (2004)

MURF1 inhibits PMA-induced translocation of PKCε from the perinuclear region to focal adhesion complexes in NRVM. NRVM were infected with Ad.GFP (A and B) or Ad.MURF1 (C and D) for 24 h in serum-free medium followed by induction with PMA (B and D) for 15 min. The cells were fixed and incubated with anti-vinculin (blue) and anti-PKCε (red) for 2 h. Arrows indicate focal adhesions.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172633&req=5

fig4: MURF1 inhibits PMA-induced translocation of PKCε from the perinuclear region to focal adhesion complexes in NRVM. NRVM were infected with Ad.GFP (A and B) or Ad.MURF1 (C and D) for 24 h in serum-free medium followed by induction with PMA (B and D) for 15 min. The cells were fixed and incubated with anti-vinculin (blue) and anti-PKCε (red) for 2 h. Arrows indicate focal adhesions.
Mentions: Because RACK1 is known to regulate targeting of PKC isoforms, we examined the translocation of PKC isoforms in NRVM in the presence of MURF1. In nonstimulated myocytes, PKCβII is associated with fibrillar structures and after activation it translocates to the nuclear and cell periphery, whereas before stimulation, PKCε is observed in the nucleus and perinucleus but translocates to cross-striated structures and focal adhesions in stimulated cells (Disatnik et al., 1994a; Ron et al., 1995; Johnson et al., 1996; Heidkamp et al., 2003). PKC activation induces the association and colocalization of RACK1 with PKCβII (Ron et al., 1999); MuRF1 modestly increased perinuclear localization of PKCβII under unstimulated conditions and did not inhibit PE- or PMA-dependent PKCβII translocation (Fig. 3). PKCβII moved from cytosol to perinuclear structures after activation with PE and PMA, and colocalized with RACK1 in control as well as Ad.MURF1-infected NRVM. Several recent reports have also shown colocalization of PKCε and RACK1 in response to stimulation with PE and PMA (Pass et al., 2001a,b; Besson et al., 2002). Using an anti-PKCε antibody, we observed the translocation of PKCε from perinuclear structures to focal adhesions upon activation with PE or PMA in NRVM infected with Ad.GFP, as indicated by costaining with the focal adhesion marker vinculin. Surprisingly, this translocation was inhibited after infection with Ad.MURF1 (Fig. 4). PKCε translocation occurred in 22 ± 6% of Ad.MURF1-infected cells treated with PMA compared with 82 ± 8% of control cells. After stimulation with PMA, PKCε was still observed in the perinuclear structures and nuclei in these cells. Similar patterns were observed after stimulation with PE (unpublished data). These data indicate that MURF1 specifically inhibits the movement of the PKCε isoform after agonist-induced stimulation.

Bottom Line: Much effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy.MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1.MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size.

View Article: PubMed Central - PubMed

Affiliation: Carolina Cardiovascular Biology Center, University of North Carolina, Chapel Hill, NC 27599, USA.

ABSTRACT
Much effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy. In spite of this, we still know little about the mechanisms that inhibit hypertrophic growth. We define a novel anti-hypertrophic signaling pathway regulated by muscle ring finger protein-1 (MURF1) that inhibits the agonist-stimulated PKC-mediated signaling response in neonatal rat ventricular myocytes. MURF1 interacts with receptor for activated protein kinase C (RACK1) and colocalizes with RACK1 after activation with phenylephrine or PMA. Coincident with this agonist-stimulated interaction, MURF1 blocks PKCepsilon translocation to focal adhesions, which is a critical event in the hypertrophic signaling cascade. MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1. MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size. These findings establish that MURF1 is a key regulator of the PKC-dependent hypertrophic response and can blunt cardiomyocyte hypertrophy, which may have important implications in the pathophysiology of clinical cardiac hypertrophy.

Show MeSH
Related in: MedlinePlus