Publication Date

2013-07-08

Availability

Embargoed

Embargo Period

2015-07-08

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Biochemistry and Molecular Biology (Medicine)

Date of Defense

2013-04-30

First Committee Member

Michael Kapiloff

Second Committee Member

Sandra Lemmon

Third Committee Member

Joy Lincoln

Fourth Committee Member

Steven Lipshultz

Fifth Committee Member

Joshua Hare

Abstract

ERK MAP-kinases regulate the concentric growth and survival of cardiac myocytes. Recently, we discovered that the ERK effector RSK3 binds to the scaffold protein mAKAP at the cardiac myocyte nuclear envelope. RSK3 activation was specifically regulated by ERK1/2/5, but not JNK and p38, in phenylephrine (PE), leukemia inhibitory factor and serum-treated neonatal cardiac myocytes. RSK3 siRNA inhibited PE-increased myocyte cross-section area 36 +/- 6%, while completely inhibiting ANF and BNP expression. To address the importance in RSK3 anchoring in its function, we expressed the RSK3 binding domain of mAKAP in myocytes. The peptide inhibited morphologic hypertrophy 63 +/- 16% and ANF expression completely. We tested whether RSK3 is required for compensated cardiac hypertrophy by subjecting the RSK3-/- mice to pressure overload for two weeks. Post-mortem gravimetric analysis showed that the corresponding increase in biventricular weight following TAC was similarly diminished in the knock-out mice (48% for RSK3+/+ vs. 26% for RSK3-/- mice). TAC primarily induces concentric growth of cardiac myocytes. Inspection of wheat germ agglutinin–stained heart sections revealed that consistent with these results, RSK3 knock-out attenuated the TAC-induced increase in myocyte transverse cross-section area by ~46%. Proportional results were obtained by morphometric analysis of adult cardiac myocytes isolated from the TAC mice. In order to characterize the RSK3-/- cardiac phenotype at a molecular level, we surveyed for differences in the cardiac expression of 30 genes encoding proteins involved either in cardiac remodeling or hypertrophic signaling. About 2/3 of the genes in our panel were significantly increased or decreased in expression by TAC. In general, the changes in expression were attenuated by RSK3 knock-out. For example, TAC-induced ANF expression was dramatically inhibited in RSK3-/- mice, consistent with the results obtained for PE-treated neonatal myocytes. Although after two weeks of pressure overload the small increases in cellular apoptosis and interstitial fibrosis detectable by histology. Interestingly, two genetic markers of fibrosis that were significantly induced in TAC wildtype mice, transforming growth factor β2 and collagen VIα1, (Yang et al., 2012) were attenuated in expression by RSK3 knock-out. Additionally, mAKAP was conditionally knocked out from cardiac myocytes in adult mice. mAKAP cardiac knock-out mice were subjected to TAC and displayed a similar attenuation of hypertrophy compared to controls. Together, these results show that mAKAP and its anchored RSK3 effector are important for concentric myocyte hypertrophy.

Keywords

Heart; Hypertrophy; mAKAP; RSK3; Cardiac; Remodeling

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