Hideo Mizutani

New Isoform of Cardiac Myosin Light Chain Kinase and the Role of Cardiac Myosin Phosphorylation in α1-Adrenoceptor Mediated Inotropic Response.

Background & Aims Cardiac myosin light chain kinase (cMLCK) plays an obligatory role in maintaining the phosphorylation levels of regulatory myosin light chain (MLC2), which is thought to be crucial for regulation of cardiac function. To test this hypothesis, the role played by ventricular MLC2 (MLC2v) phosphorylation was investigated in the phenylephrine-induced increase in twitch tension using the naturally-occurring mouse strain, C57BL/6N, in which cMLCK is down regulated. Methods and Results By Western blot and nanoLC-MS/MS analysis, cMLCKs with molecular mass of 61-kDa (cMLCK-2) and/or 86-kDa were identified in mice heart. Among various mouse strains, C57BL/6N expressed cMLCK-2 alone and the closest relative strain C57BL/6J expressed both cMLCKs. The levels of MLC2v phosphorylation was significantly lower in C57BL/6N than in C57BL/6J. The papillary muscle twitch tension induced by electrical field stimulation was smaller in C57BL/6N than C57BL/6J. Phenylephrine had no effect on MLC2v phosphorylation in either strains but increased the twitch tension more potently in C57BL/6J than in C57BL/6N. Calyculin A increased papillary muscle MLC2v phosphorylation to a similar extent in both strains but increased the phenylephrine-induced inotropic response only in C57BL/6N. There was a significant positive correlation between the phenylephrine-induced inotropic response and the levels of MLC2v phosphorylation within ranges of 15–30%. Conclusions We identified a new isoform of cMLCK with a molecular mass of 61kDa(cMLCK-2) in mouse heart. In the C57BL/6N strain, only cMLCK-2 was expressed and the basal MLC2v phosphorylation levels and the phenylephrine-induced inotropic response were both smaller. We suggest that a lower phenylephrine-induced inotropic response may be caused by the lower basal MLC2v phosphorylation levels in this strain.

Big mitogen-activated protein kinase: a new player in vascular remodeling.

Big mitogen-activated protein kinase (BMK1), also termed extracellular signal regulated kinase 5 (ERK5), is the most recently identified mitogen-activated protein (MAP) kinase family member, and has twice the molecular mass of other MAP kinases. Similar to ERK1/ERK2, the N-terminal half of BMK1 contains both a kinase catalytic domain and a TEY activation motif, while its C-terminal tail is unique and is involved in BMK1-specific molecular functions, including the regulation of the transcription activity of target molecules (1, 2). As in the cases of the other MAP kinases, BMK1 is activated in response to a variety of stimuli, namely oxidative stress, hyperosmolarity, serum and growth factors (1). In response to these stimuli, the BMK1 cascade is activated by sequentially activated kinases, which consist of the kinases MEKK2/MEKK3, MEK5, and then BMK1 (1, 2). BMK1 signaling plays roles not only in physiological cellular responses, including cell proliferation, differentiation, and survival, but also under pathological conditions such as carcinogenesis and cardiovascular diseases (1). In a genetic model of hypertension and congestive heart failure, BMK1 activity was shown to be increased during left ventricular hypertrophy and then reverted to normal during the period of congestive heart failure (3). Overexpression of the constitutively active form of MEK5, a highly specific upstream kinase for ERK5, induced cardiomyocyte elongation by interfering with parallel assembly of sarcomeres in vitro (4). Furthermore, cardiac-specific overexpression of activated MEK5 in mice resulted in rapidly decompensating eccentric cardiac hypertrophy and sudden death, indicating that MEK5-BMK1 plays a role in the induction of eccentric cardiac hypertrophy in vivo (4). In contrast, BMK1 has also been reported to play a protective role in ischemia/perfusioninduced cardiac injury (5). BMK1 is activated in endothelial cells (EC) by shear stress (6), and activation of BMK1 protects EC from apoptosis (7), indicating that the action of BMK1 in EC is atheroprotective. Indeed, in recent genetic studies, endothelial-specific knockout mice demonstrated embryonic lethality due to the effect of BMK1 knockout on endothelial function and resultant impaired blood vessel integrity, suggesting that the BMK1 pathway is essential for endothelial survival and the maintenance of blood vessel integrity in vivo (8). Platelet-derived growth factor (PDGF) is recognized as a major mitogen in serum and one of the most important growth factors promoting the proliferation of vascular smooth muscle cells (VSMC) and atherogenesis (9). The MAP kinases including ERK, c-Jun N-terminal kinase (JNK) and p38 had been reported to participate in both PDGF-induced VSMC proliferation and migration (10). Although BMK1 was initially reported to be poorly activated in rat VSMC following stimulation with PDGF (11), several recent reports have indicated that stimulation with aldosterone (12) and growth factors including PDGF and fibroblast growth factor-2 (FGF-2) (13, 14) do activate BMK1, which then plays a role in VSMC proliferation. Migration of VSMC is one of the essential processes in the progression of atherosclerosis. In an article appearing in this issue of Hypertension Research, Izawa et al. demonstrate a new role of BMK1 in VSMC migration (15). Namely, their