Calcineurin Promotes Protein Kinase C and c-Jun NH2-terminal Kinase Activation in the Heart

doi:10.1074/jbc.275.18.13571

Calcineurin Promotes Protein Kinase C and c-Jun NH₂-terminal Kinase Activation in the Heart

CROSS-TALK BETWEEN CARDIAC HYPERTROPHIC SIGNALING PATHWAYS*

From the ^‡Department of Pediatrics, University of Cincinnati, Children's Hospital Medical Center, Cincinnati, Ohio 45229-3039 and the ^¶Department of Medicine, Cardiovascular Research Center, Massachusetts General Hospital, Harvard School of Medicine, Charleston, Massachusetts 02129-2060

Abstract

Multiple intracellular signaling pathways have been shown to regulate the hypertrophic growth of cardiomyocytes. Both necessary and sufficient roles have been described for the mitogen activated protein kinase¹ (MAPK) signaling pathway, specific protein kinase C (PKC) isoforms, and calcineurin. Here we investigate the interdependence between calcineurin, MAPK, and PKC isoforms in regulating cardiomyocyte hypertrophy using three separate approaches. Hearts from hypertrophic calcineurin transgenic mice were characterized for PKC and MAPK activation. Transgenic hearts demonstrated activation of c-Jun NH₂-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK1/2), but not p38 MAPK factors. Calcineurin transgenic hearts demonstrated increased activation of PKCα, β₁, and θ, but not of ε, β₂, or λ. In a second approach, cultured cardiomyocytes were infected with a calcineurin adenovirus to induce hypertrophy and the effects of pharmacologic inhibitors or co-infection with a dominant negative adenovirus were examined. Calcineurin-mediated hypertrophy was prevented with PKC inhibitors, Ca²⁺chelation, and attenuated with a dominant negative SEK-1 (MKK4) adenovirus, but inhibitors of ERK or p38 activation had no effect. In a third approach, we examined the activation of MAPK factors and PKC isoforms during the progression of load-induced hypertrophy in aortic banded rats with or without cyclosporine. We determined that inhibition of calcineurin activity with cyclosporine prevented PKCα, θ, and JNK activation, but did not affect PKCε, β, λ, ERK1/2, or p38 activation. Collectively, these data indicate that calcineurin hypertrophic signaling is interconnected with PKCα, θ, and JNK in the heart, while PKCε, β, λ, p38, and ERK1/2 are not involved in calcineurin-mediated hypertrophy.

Footnotes

↵* This work was supported in part by National Institutes of Health Grants HL69562 and HL-62927 (to J. D. M) and DK50282 and HL61688 (to T. F.), the Pew charitable trust foundation (to J. D. M.), an American Heart Association grant-in-aid (to J. D. M.), and a post-doctoral award (to L. J. D. W.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵§ Contributed equally to the results of this study.
↵‖ Established Investigator of the American Heart Association.
↵** Pew Scholar. To whom correspondence should be addressed:Div. of Molecular Cardiovascular Biology, Dept. of Pediatrics, Children's Hospital Medical Center, 3333 Burnet Ave., Cincinnati, OH 45229-3039. E-mail:molkj0@chmcc.org.
↵2 A. Yatani, J. D. Molkentin, and E. G. Kranias, unpublished observation.
Abbreviations:

ERK1/2

extracellular signal-regulated kinase 1/2

MAPK

mitogen-activated protein kinase

MKK

MAP kinase kinase

PKC

protein kinase C

JNK

c-Jun NH₂-terminal kinase

ANF

atrial natriuretic factor

BAPTA-AM

1,2-bis(2-amino-phenoxy)ethane-N,N,N′,N′-tetraacetic acid-acetoxymethylester

PMA

phorbol 12-myristate 13-acetate

AdCnA

calcineurin adenovirus

Adβgal

β-galactosidase adenovirus

ECF

enhanced chemiflorescence

ANOVA

analysis of variance

PP2A or B

protein phosphatase 2A or B

PE

phenylephrine

BDM

butanedione monoxime

TG

transgenic
- Received November 12, 1999.
- Revision received February 11, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.