Modulating protein kinase D1 signal transduction

Protein kinase C （PKC） consists of a family of serine/threonine kinases that are identified by the presence of two copies of the C1 domain, which form the diacylglycerol （DAG）-binding module. According to their enzymatic activities PKCs are sub-divided into conventional isozymes （PKCα, β and γ; calcium, phospholipid and DAG-activated kinases）, novel isozymes （PKCδ, ε, η, μ and θ; calcium-insensitive, phospholipid-dependent and DAG-dependent）, and atypical isozymes （PKCζ and λ; calcium-insensitive and DAG-insensitive enzymes）. Human protein kinase Cμ and its mouse homolog, protein kinase D1 （PKD1）, which has been under intense investigation in recent years, is a DAG-dependent, Ca^2＋-independent serine/threonine protein kinase as a novel PKC isoform. Recently PKDs were classified as a novel subgroup of the calcium/calmodulin-dependent protein kinase （CAMK） family, based on sequence similarities of the kinase domain; the catal^ctic domain of PKD1 has the highest homology to CAMK. PKD1 has three main pathways for activation. One is DAG-phospholipase C （PLC）-PKC-dependent activation of PKD1. In this model, PKD 1 not only acts as a direct DAG target but also lies downstream of PKCs to regulate biological processes in cells.

Cyclovirobuxinum D Alleviates Cardiac Hypertrophy in Hyperthyroid Rats by Preventing Apoptosis of Cardiac Cells and Inhibiting the p38 Mitogen-Activated Protein Kinase Signaling Pathway

Objective： To investigate the underlying mechanisms of cyclovirobuxinum D（Cvb-D） on alleviating cardiac hypertrophy in rats. Methods： Sprague-Dawley rats were randomly divided into 5 groups： control group; levothyroxine-induced cardiac hypertrophy group（model）; levothyroxine-induced cardiac hypertrophy ＋ Cvb-D group（Cvb-D）; levothyroxine-induced cardiac hypertrophy ＋ captopril group（captopril）; levothyroxine-induced cardiac hypertrophy ＋ SB203580 group（SB203580）, n=10 for each group. Rats were daily administered the respective drugs continuously for14 days by gastric gavage. A rat model of cardiac hypertrophy was established by intraperitoneal injection of levothyroxine to investigate whether Cvb-D protects against cardiac hypertrophy by inhibiting the p38 mitogen-activated protein kinase（MAPK） signaling pathway and preventing apoptosis of cardiac cells. Results： Treatment with Cvb-D significantly deceased left ventricle hypertrophy, improved the histopathology, hemodynamic conditions, and cardiac function in rats with cardiac hypertrophy. Compared with the normal control group, in rats with cardiac hypertrophy, expression of bax in the heart and phospho-p38 MAPK protein levels were significantly up-regulated（P〈0.01 or 0.05）, whereas the bcl-2 protein level was downregulated（P〈0.01）. In contrast, Cvb-D treatment reversed the changes in bax and phospho-p38 MAPK protein levels but increased the bcl-2 protein level（P〈0.01 or 0.05）, and these effects were similar to those of captopril and SB203580（a specific p38 MAPK inhibitor） treatment. Furthermore, both Cvb-D, captopril and SB203580 reduced m RNA expression of p38α, p38β, c-fos, and c-jun m RNA, and Cvb-D had a stronger effect（P〈0.01）. Conclusion： These results demonstrate that Cvb-D protects against cardiac hypertrophy, which is possibly mediated by prevention of cardiac cell apoptosis and inhibition of the p38 MAPK signaling pathway.

Disruption of dopamine D1 receptor phosphorylation at serine 421 attenuates cocaine-induced behaviors in mice

Dopamine D1 receptors（D1Rs） play a key role in cocaine addiction, and multiple protein kinases such as GRKs, PKA, and PKC are involved in their phosphorylation. Recently, we reported that protein kinase D1 phosphorylates the D1 R at S421 and promotes its membrane localization. Moreover, this phosphorylation of S421 is required for cocaineinduced behaviors in rats. In the present study, we generated transgenic mice over-expressing S421A-D1 R in the forebrain. These transgenic mice showed reduced phospho-D1R（S421） and its membrane localization, and reduced downstream ERK1/2 activation in the striatum. Importantly, acute and chronic cocaine-induced locomotor hyperactivity and conditioned place preference were significantly attenuated in these mice. These findings provide in vivo evidence for the critical role of S421 phosphorylation of the D1 R in its membrane localization and in cocaine-induced behaviors. Thus, S421 on the D1 R represents a potential pharmacotherapeutic target for cocaine addiction and other drug-abuse disorders.