Lysyl-tRNA synthetase (LysRS) was found to produce diadenosine tetraphosphate (Ap(4)A) in vitro more than two decades ago. Here, we used LysRS silencing in mast cells in combination with transfected normal and mutated LysRS to demonstrate in vivo the critical role played by LysRS in the production of Ap(4)A in response to immunological challenge. Upon such challenge, LysRS was phosphorylated on serine 207 in a MAPK-dependent manner, released from the multisynthetase complex, and translocated into the nucleus. We previously demonstrated that LysRS forms a complex with MITF and its repressor Hint-1, which is released from the complex by its binding to Ap(4)A, enabling MITF to transcribe its target genes. Here, silencing LysRS led to reduced Ap(4)A production in immunologically activated cells, which resulted in a lower level of MITF inducible genes. Our data demonstrate that specific LysRS serine 207 phosphorylation regulates Ap(4)A production in immunologically stimulated mast cells, thus implying that LysRS is a key mediator in gene regulation.

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The antigen-mediated activation of mast cells by means of IgE antibodies bound to the cell surface leads to direct interactions between FcɛRI receptor cytoplasmic domains and various intracellular proteins. These interactions initiate diverse signal-transduction pathways, and the activation of these pathways results in the immediate release of proinflammatory agents. A delayed response also occurs and includes the release of various cytokines. It is clear that the activation of kinases is a requirement for the exocytosis observed in mast cells. In addition to the tyrosine phosphorylation of the affected system by soluble tyrosine kinases, activity of protein kinase C (PKC) results in serine or threonine phosphorylation of multiple protein substrates. In this study, we found that mast cells derived from PKCβ-deficient mice produce less interleukin 6 in response to IgE-Ag. The inhibition of exocytosis in the PKCβ-deficient mast cells occurred whether the stimuli were due to the aggregation of the mast cell surface FcɛRI or to the calcium ionophore, ionomycin. However, no significant changes were observed in the proliferative response of the mast cells to interleukin 3 (IL-3) or in their apoptotic rate after IL-3 depletion.

We previously discovered that microphthalmia transcription factor (MITF) and upstream stimulatory factor 2 (USF2) each forms a complex with its inhibitor histidine triad nucleotide-binding 1 (Hint-1) and with lysyl-tRNA synthetase (LysRS). Moreover, we showed that the dinucleotide diadenosine tetraphosphate (

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ErbB2 interacting protein (Erbin) is a widely expressed protein and participates in inhibition of several intracellular signaling pathways. Its mRNA has been found to be present in relatively high levels in the heart. However, its physiological role in the heart has not been explored. In the present work, we elucidated the role of Erbin in cardiac hypertrophy. Cardiac hypertrophy was induced in mice either by isoproterenol administration or by aortic constriction. The level of Erbin was significantly decreased in both models. Erbin −/− mice rapidly develop decompensated cardiac hypertrophy, and following severe pressure overload all Erbin −/− mice died from heart failure. Down-regulation of Erbin expression was also observed in biopsies derived from human failing hearts. It is known that Erbin inhibits Ras-mediated activation of the extracellular signal-regulated kinase (ERK) by binding to Soc-2 suppressor of clear homolog (Shoc2). Our data clearly show that ERK phosphorylation is enhanced in the heart tissues of Erbin −/− mice. Furthermore, we clearly demonstrate here that Erbin associates with Shoc2 in both whole hearts and in cardiomyocytes, and that in the absence of Erbin, Raf is phosphorylated and binds Shoc2, resulting in ERK phosphorylation. In conclusion, Erbin is an inhibitor of pathological cardiac hypertrophy, and this inhibition is mediated, at least in part, by modulating ERK signaling.C ardiovascular disease remains the number one cause of mortality in the Western world, with heart failure representing the fastest-growing subclass over the past decade (1, 2). In myocardial hypertrophy caused by exercise, pregnancy, or developmental signals, cardiac structure and function are normal. However, in response to insults such as sustained excessive workloads, pathological cardiac hypertrophy is characterized by changes in contractility, loss of myocytes with fibrosis, systolic or diastolic dysfunction, and fetal gene reactivation. Pathological cardiac hypertrophy predisposes individuals to heart failure, arrhythmia, and sudden death (1, 2). The tyrosine kinase receptor avian erythroblastic leukemia viral oncogene homolog 2 (ErbB2), also known in humans as Her2, is an important regulator in cardiac hypertrophy development and heart failure (3). ErbB2-deficient conditional mutants develop dilated cardiomyopathy (3). ErbB2 signaling in cardiomyocytes is therefore essential for the prevention of dilated cardiomyopathy. However, the mechanism by which ErbB2 exerts its cardiac effects in general, and on cardiac hypertrophy in particular, is poorly defined.ErbB2 interacting protein (Erbin), a member of the leucine-rich repeat and PDZ domain proteins (4), was originally reported as a binding partner of Her2/neu (ErbB2) (4). Erbin is a widely expressed protein, and high levels of Erbin are present in the heart (5). Recently, Erbin has been demonstrated to form a complex with Her2 and β2-adrenergic receptor in cardiomyocytes. Also, Erbin has been shown to protect cardiomyocytes from apoptosis induced by chronic ca...