N-myristoylated proteins, key components in intracellular signal transduction systems enabling rapid and flexible cell responses

N-myristoylation, one of the co- or post-translational modifications of proteins, has so far been regarded as necessary for anchoring of proteins to membranes. Recently, we have revealed that Nα-myristoylation of several brain proteins unambiguously regulates certain protein–protein interactions that may affect signaling pathways in brain. Comparison of the amino acid sequences of myristoylated proteins including those in other organs suggests that this regulation is involved in signaling pathways not only in brain but also in other organs. Thus, it has been shown that myristoylated proteins in cells regulate the signal transduction between membranes and cytoplasmic fractions. An algorithm we have developed to identify myristoylated proteins in cells predicts the presence of hundreds of myristoylated proteins. Interestingly, a large portion of the myristoylated proteins thought to take part in signal transduction between membranes and cytoplasmic fractions are included in the predicted myristoylated proteins. If the proteins functionally regulated by myristoylation, a posttranslational protein modification, were understood as cross-talk points within the intracellular signal transduction system, known signaling pathways could thus be linked to each other, and a novel map of this intracellular network could be constructed. On the basis of our recent results, this review will highlight the multifunctional aspects of protein N-myristoylation in brain

RG100204, A Novel Aquaporin-9 Inhibitor, Reduces Septic Cardiomyopathy and Multiple Organ Failure in Murine Sepsis

International audienceSepsis is caused by systemic infection and is a major health concern as it is the primary cause of death from infection. It is the leading cause of mortality worldwide and there are no specific effective treatments for sepsis. Gene deletion of the neutral solute channel Aquaporin 9 (AQP9) normalizes oxidative stress and improves survival in a bacterial endotoxin induced mouse model of sepsis. In this study we described the initial characterization and effects of a novel small molecule AQP9 inhibitor, RG100204, in a cecal ligation and puncture (CLP) induced model of polymicrobial infection. In vitro, RG100204 blocked mouse AQP9 H 2 O 2 permeability in an ectopic CHO cell expression system and abolished the LPS induced increase in superoxide anion and nitric oxide in FaO hepatoma cells. Pre-treatment of CLP-mice with RG100204 (25 mg/kg p.o. before CLP and then again at 8 h after CLP) attenuated the hypothermia, cardiac dysfunction (systolic and diastolic), renal dysfunction and hepatocellular injury caused by CLP-induced sepsis. Post-treatment of CLP-mice with RG100204 also attenuated the cardiac dysfunction (systolic and diastolic), the renal dysfunction caused by CLP-induced sepsis, but did not significantly reduce the liver injury or hypothermia. The most striking finding was that oral administration of RG100204 as late as 3 h after the onset of polymicrobial sepsis attenuated the cardiac and renal dysfunction caused by severe sepsis. Immunoblot quantification demonstrated that RG100204 reduced activation of the NLRP3 inflammasome pathway. Moreover, myeloperoxidase activity in RG100204 treated lung tissue was reduced. Together these results indicate that AQP9 may be a novel drug target in polymicrobial sepsis

N-terminal domain linkage modulates the folding properties of protein S epidermal growth factor-like modules

Protein S interacts with activated protein C to play a crucial role in blood anticoagulation, and protein S deficiency is associated with increased risk of thrombosis. Despite the large volume of functional data available for this protein, no atomic resolution structure data have yet been reported. This is due at least in part to difficulties encountered when trying to produce fragments dissected from the intact protein; however, a few successful strategies have been described. In this research we have expressed a number of constructs containing protein S epidermal growth factor-like (EGF) domains 1 and 2 in Escherichia coli and Pichia pastoris. None of the proteins produced was stably folded as assayed by solution nuclear magnetic resonance spectroscopy. We therefore constructed a series of non-native protein S EGF concatemers to investigate the role of pairwise domain linkage in domain folding. Our results demonstrate that N-terminal domain linkage can either positively or negatively impact on the refolding of an adjacent domain. Furthermore, analysis of the NMR data for EGF3-4 reveals the expected interdomain NOEs that are characteristic of an extended arrangement of calcium-binding EGF domains and a similar average [1H]-15N heteronuclear NOE value for each of the two domains. These results provide the first data in support of protein S EGF3-4 adopting the same extended domain orientation as observed for the functionally distinct proteins fibrillin-1 and the low-density lipoprotein receptor. The results also have important implications for future studies, particularly when a dissection approach is used, of tandem EGF domains from protein S and other proteins