Dysregulation of suppressor of cytokine signaling 3 in keratinocytes causes skin inflammation mediated by interleukin-20 receptor-related cytokines

Homeostatic regulation of epidermal keratinocytes is controlled by the local cytokine milieu. However, a role for suppressor of cytokine signaling (SOCS), a negative feedback regulator of cytokine networks, in skin homeostasis remains unclear. Keratinocyte specific deletion of Socs3 (Socs3 cKO) caused severe skin inflammation with hyper-production of IgE, epidermal hyperplasia, and S100A8/9 expression, although Socs1 deletion caused no inflammation. The inflamed skin showed constitutive STAT3 activation and up-regulation of IL-6 and IL-20 receptor (IL-20R) related cytokines, IL-19, IL-20 and IL-24. Disease development was rescued by deletion of the Il6 gene, but not by the deletion of Il23, Il4r, or Rag1 genes. The expression of IL-6 in Socs3 cKO keratinocytes increased expression of IL-20R-related cytokines that further facilitated STAT3 hyperactivation, epidermal hyperplasia and neutrophilia. These results demonstrate that skin homeostasis is strictly regulated by the IL-6-STAT3-SOCS3 axis. Moreover, the SOCS3-mediated negative feedback loop in keratinocytes has a critical mechanistic role in the prevention of skin inflammation caused by hyperactivation of STAT3.     

Disruption of Sept6, a fusion partner gene of MLL, does not affect ontogeny, leukemogenesis induced by MLL-SEPT6, or phenotype induced by the loss of Sept4

Septins are evolutionarily conserved GTP-binding proteins that can heteropolymerize into filaments. Recent studies have revealed that septins are involved in not only diverse normal cellular processes but also the pathogenesis of various diseases, including cancer. SEPT6 is ubiquitously expressed in tissues and one of the fusion partner genes of MLL in the 11q23 translocations implicated in acute leukemia. However, the roles of this septin in vivo remain elusive. We have developed Sept6-deficient mice that exhibited neither gross abnormalities, changes in cytokinesis, nor spontaneous malignancy. Sept6 deficiency did not cause any quantitative changes in any of the septins evaluated in this study, nor did it cause any additional changes in the Sept4-deficient mice. Even the depletion of Sept11, a close homolog of Sept6, did not affect the Sept6-null cells in vitro, thus implying a high degree of redundancy in the septin system. Furthermore, a loss of Sept6 did not alter the phenotype of myeloproliferative disease induced by MLL-SEPT6, thus suggesting that Sept6 does not function as a tumor suppressor. To our knowledge, this is the first report demonstrating that a disruption of the translocation partner gene of MLL in 11q23 translocation does not contribute to leukemogenesis by the MLL fusion gene.     

Biochemical and physiological evidence that calmodulin is involved in the taste response of the sugar receptor cells of the blowfly, Phormia regina

The gustatory system is essential for almost all animals. However, the signal transduction mechanisms have not yet been fully elucidated. We isolated labellar chemosensilla from blowfly, Phormia regina, and purified calcium binding proteins from the water soluble fraction. The most abundant calcium-binding protein was calmodulin. To investigate the role of calmodulin in taste transduction, electrophysiological responses were recorded with the calmodulin inhibitor, W-7. When we stimulated the labellar chemosensillum with sucrose plus W-7, a dose-dependent decrease of impulse frequency was observed when the concentration was <50 microM. In addition, when W-7 at 50 microM or higher concentration was added, an initial short-term impulse generation from the sugar receptor cell was observed, but this was followed by a silent period. When the sensillum was stimulated with W-7 plus a membrane-permeable cGMP analog, dibtyryl-cGMP or 8-bromo-cGMP, impulses of the sugar receptor cell were induced but the frequency was decreased. By the sidewall-recording method, we observed that the receptor potential induced by sucrose stimulation was decreased by W-7 in the sugar receptor cell, and corresponded with a disappearance of impulses. These data strongly suggest that the cGMP-gated channel generating receptor potential in the sugar receptor cell requires calmodulin for its gating.     

Prediction and validation of the distinct dynamics of transient and sustained ERK activation

To elucidate the hidden dynamics of extracellular-signal-regulated kinase (ERK) signalling networks, we developed a simulation model of ERK signalling networks by constraining in silico dynamics based on in vivo dynamics in PC12 cells. We predicted and validated that transient ERK activation depends on rapid increases of epidermal growth factor and nerve growth factor (NGF) but not on their final concentrations, whereas sustained ERK activation depends on the final concentration of NGF but not on the temporal rate of increase. These ERK dynamics depend on Ras and Rap1 dynamics, the inactivation processes of which are growth-factor-dependent and -independent, respectively. Therefore, the Ras and Rap1 systems capture the temporal rate and concentration of growth factors, and encode these distinct physical properties into transient and sustained ERK activation, respectively.     

Actin filaments play an essential role for transport of nascent HIV-1 proteins in host cells

To investigate the role of actin filaments (F-actin) for human immunodeficiency virus type 1 (HIV-1) production in host cells, the effect of mycalolide B that is a novel actin-depolymerizing marine toxin was examined. Mycalolide B blocked the production of HIV-1 from primary infected T-lymphoblastoid and clonically infected monocytoid cells in a concentration-dependent manner. In the presence of 10 microM of mycalolide B, F-actins were disorganized and mostly disappeared in the host cells, and viral envelope- and capsid-proteins did not reach the plasma membrane, but were distributed in the cytoplasm forming aggregates. In electron micrographs, no HIV-1 virions were detected on the cell surface, but many lysosome-like vesicles containing electron dense granules were observed in the cytoplasm, implying that mycalolide B did not disturb the synthesis of viral proteins, but rather inhibited their transport processes of HIV-1 in the host cells.     

Molecular mechanism of changes in the morphine-induced pharmacological actions under chronic pain-like state: suppression of dopaminergic transmission in the brain

In the present study, we demonstrated whether a neuropathic pain-like state induced by sciatic nerve ligation in rodents could cause a long-lasting change in intracellular signaling in both supraspinal and spinal cord related to the suppression of morphine's effect. Mice with sciatic nerve ligation exhibited a significant suppression of the morphine-induced antinociception. Under this condition, phosphorylated-conventional protein kinase C-like immunoreactivity (p-cPKC-IR) and phosphorylated-micro-opioid receptor (p-MOR)-IR were clearly increased on the ipsilateral side in the dorsal horn of the spinal cord of nerve-ligated mice. It is of interest to note that astroglial hypertrophy as well as its proliferation was also noted in this area of sciatic nerve-ligated mice. Like nerve injury, the increase in cPKC activities and astroglial hypertrophy/proliferation in this region was observed by repeated morphine treatment. These findings suggest that the phosphorylation of both cPKC and MOR in the dorsal horn of the spinal cord by sciatic nerve ligation may play a substantial role in the suppression of morphine-induced antinociception under a neuropathic pain-like state. Sciatic nerve injury also caused a significant inhibition of MOR-mediated G-protein activation onto GABAergic neurons and a dramatic reduction in ERK activities onto dopaminergic neurons in the ventral tegmental area (VTA) regulating the rewarding effect of opioids. Furthermore, we found that the inhibition of ERK cascade in the VTA by treatment with specific inhibitors suppressed the morphine-induced rewarding effect in normal mice. These findings provide evidence that the direct reduction in MOR function and the persistent decrease in ERK activity of dopaminergic neurons in the VTA may contribute to the suppression of the morphine-induced rewarding effect under a neuropathic pain-like state. Conclusively, our recent findings provide novel evidences for the mechanism underlying the less sensitivity to opioids under a neuropathic pain-like state.     

6-Carboxy-5,7-diarylcyclopenteno[1,2-b]pyridine derivatives: a novel class of endothelin receptor antagonists

Compounds (2-5) with a 6-carboxy-5,7-diarylcyclopentenopyridine skeleton were designed, synthesized, and identified as a new class of potent non-peptide endothelin receptor antagonists. The regio-isomer 2 was found to show potent inhibitory activity with an IC(50) value of 2.4 nM against (125)I-labeled ET-1 binding to human ET(A) receptors and a 170-fold selectivity for ET(A) over ET(B) receptors. Furthermore, 2 displayed more potent in vivo activity than did the indan-type compound 1 in a mouse ET-1 induced lethality model, suggesting the potential of 2 as a new lead structure. Derivatization on substituted phenyl groups at the 5- and 7-positions of 2 revealed that a 3,4-methylenedioxyphenyl group at the 5-position and a 4-methoxyphenyl group at the 7-position were optimal for binding affinity. Further derivatization of 2 by incorporating a substituent into the 2-position of the 4-methoxyphenyl group led to the identification of a more potent ET(A) selective antagonist 2p with an IC(50) value of 0.87 nM for ET(A) receptors and a 470-fold selectivity. In addition, 2p showed highly potent in vivo efficacy (AD(50): 0.04 mg/kg) in the lethality model.