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Identification of protein kinase G I alpha interacting proteins as potential targets to prevent cardiac remodeling


We recently reported that mutation of the cGMP-dependent Protein Kinase G I alpha (PKGIα) N-terminal leucine zipper (LZ) domain (in the PKGIα LZ mutant, or LZM, mouse) accelerates cardiac remodeling and heart failure after left ventricular (LV) pressure overload, and prevents the anti-remodeling effect of sildenafil [1]. We therefore hypothesized that PKGIα attenuates remodeling by regulating cardiac signaling pathways that are dependent on substrate interactions mediated by its LZ domain. As a first step to identifying cardiac proteins downstream of PKGIα, we screened myocardial lysates for PKGIα LZ domain-interacting proteins. Our previous work revealed a requirement for the PKGIα LZ domain for the activation of anti-remodeling myocardial JNK activity after LV pressure overload. MLK3 is a MAPKKK that contains an LZ domain and activates JNK.


We now demonstrate, by immunoprecipitation, that MLK 3 interacts with the PKGIα LZ domain in myocardial lysates. We show further that 8-Br-cGMP induces MLK3 phosphorylation on Thr277 and Ser281 in WT, but not LZM lysates. And, in 293 cells transfected with FLAG-MLK3, 8Br-cGMP induced PKGIα-MLK3 co-precipitation, and increased MLK3 phosphorylation on Thr277/Ser281. Co-transfection of MLK3 and PKGIα also induced MLK3 phosphorylation. We next examined the cardiovascular effect of MLK3 deletion in vivo. Male 8 week old MLK3-/- mice display basal bi-ventricular hypertrophy compared with littermate controls (LV/Tibia length 42.8 ± 0.6 mg/cm in WT, 52.9 ± 1.8 in MLK3 -/-; P<0.01; RV/TL 10.8 ± 0.1 mg/cm in WT, 13.3 ± 0.3 in MLK3-/-; P<0.01; n= 7 WT, 5 MLK3-/-). By 14-16 weeks of age, LVH progressed in the MLK3-/- mice (LV/TL 47.7 ± 1.3 mg/cm in WT, 59.8 ± 7.5 in MLK3-/-; n= 6 WT, 9 MLK3-.-; P<0.01). Arterial blood pressure was modestly increased, though still normal, in MLK3-/- mice (SBP 93 ± 1 in WT, 113 ± 1 in MLK3-/-). And, 14-16 week MLK3-/- mice have impaired LV diastolic function (tau 3.2 ± 0.1 ms WT, 3.7 ± 0.1 MLK3-/-; P 0.06).


Our studies reveal a novel function of MLK3 as a myocardial PKGIα effector and inhibitor of LVH. These results support the strategy of exploring LZ-dependent PKGIα substrates in the myocardium to identify potential therapeutic targets for cardiac remodeling.


  1. Blanton R, Takimoto , Lane AM, Aronovitz M, Piotrowski R, Karas RH, Kass DA, Mendelsohn ME: Protein kinase g iα inhibits pressure overload-induced cardiac remodeling and is required for the cardioprotective effect of sildenafil in vivo. J Am Heart Assoc. 2012, 1: e003731-10.1161/JAHA.112.003731.

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Correspondence to Robert Blanton.

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This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Blanton, R., Lane, A., Aronovitz, M. et al. Identification of protein kinase G I alpha interacting proteins as potential targets to prevent cardiac remodeling. BMC Pharmacol Toxicol 14 (Suppl 1), P10 (2013).

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