Novel gating and sensitizing mechanism of capsaicin receptor (TRPV1): tonic inhibitory regulation of extracellular sodium through the external protonation sites on TRPV1

Transient receptor potential V1 (TRPV1) is a nonselective cation channel expressed in nociceptors and activated by capsaicin. TRPV1 detects diverse stimuli, including acid, heat, and endogenous vanilloids, and functions as a molecular integrator of pain perception. Herein we demonstrate a novel regulatory role of extracellular Na(+) ([Na(+)](o)) on TRPV1 function. In human embryonic kidney 293 cells expressing porcine TRPV1, low [Na(+)](o) evoked increases of [Ca(2+)](i) that were suppressed by TRPV1 antagonists and facilitated responses to capsaicin, protons, heat, and an endovanilloid. [Na(+)](o) removal simultaneously elicited a [Ca(2+)](i) increase and outward-rectified current with a reversal potential similar to those of capsaicin. Neutralization of the two acidic residues which confer the proton sensitivity to TRPV1 resulted in a reduction of low [Na(+)](o)-induced responses. In primary culture of porcine sensory neurons, the removal of [Na(+)](o) produced a [Ca(2+)](i) increase and current responses only in the cells responding to capsaicin. Low [Na(+)](o) evoked a [Ca(2+)](i) increase in sensory neurons of wild type mice, but not TRPV1-null mice, and in human embryonic kidney 293 cells expressing human TRPV1. The present results suggest that [Na(+)](o) negatively regulates the gating and polymodal sensitization of the TRPV1 channel. [Na(+)](o) surrounding several proton-sensitive sites on the extracellular side of the pore-forming loop of the TRPV1 channel may play an important role as a brake to suppress the excessive activity of this channel under physiological conditions.     

NMDA receptor regulates migration of newly generated neurons in the adult hippocampus via Disrupted-In-Schizophrenia 1 (DISC1)

In the mammalian brain, new neurons are continuously generated throughout life in the dentate gyrus (DG) of the hippocampus. Previous studies have established that newborn neurons migrate a short distance to be integrated into a pre-existing neuronal circuit in the hippocampus. How the migration of newborn neurons is governed by extracellular signals, however, has not been fully understood. Here, we report that NMDA receptor (NMDA-R)-mediated signaling is essential for the proper migration and positioning of newborn neurons in the DG. An intraperitoneal injection of the NMDA-R antagonists, memantine, or 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) into adult male mice caused the aberrant positioning of newborn neurons, resulting in the overextension of their migration in the DG. Interestingly, we revealed that the administration of NMDA-R antagonists leads to a decrease in the expression of Disrupted-In-Schizophrenia 1 (DISC1), a candidate susceptibility gene for major psychiatric disorders such as schizophrenia, which is also known as a critical regulator of neuronal migration in the DG. Furthermore, the overextended migration of newborn neurons induced by the NMDA-R antagonists was significantly rescued by exogenous expression of DISC1. Collectively, these results suggest that the NMDA-R signaling pathway governs the migration of newborn neurons via the regulation of DISC1 expression in the DG.     

Autoregulation and homodimerization are involved in the activation of the plant steroid receptor BRI1

The leucine-rich-repeat receptor serine/threonine kinase, BRI1, is a cell-surface receptor for brassinosteroids (BRs), the steroid hormones of plants, yet its activation mechanism is unknown. Here, we report a unique autoregulatory mechanism of BRI1 activation. Removal of BRI1's C terminus leads to a hypersensitive receptor, indicated by suppression of dwarfism of BR-deficient and BR-perception mutants and by enhanced BR signaling as a result of elevated phosphorylation of BRI1. Several sites in the C-terminal region can be phosphorylated in vitro, and transgenic Arabidopsis expressing BRI1 mutated at these sites demonstrates an essential role of phosphorylation in BRI1 activation. BRI1 is a ligand-independent homo-oligomer, as evidenced by the transphosphorylation of BRI1 kinase in vitro, the dominant-negative effect of a kinase-inactive BRI1 in transgenic Arabidopsis, and coimmunoprecipitation experiments. Our results support a BRI1-activation model that involves inhibition of kinase activity by its C-terminal domain, which is relieved upon ligand binding to the extracellular domain.     

Cloning and Characterization of the cDNA Encoding a Novel Brain-Specific 14-kDa Protein

A new acidic protein with a molecular weight of 14,000 was purified from rat brain, in which it was specifically expressed, and partially sequenced by protein sequencing. On the basis of results obtained from the amino acid sequences, mixed oligonucleotides were synthesized and used as probes to clone a cDNA from a rat brain cDNA library. The cloned cDNA provided the full-length sequence of the 14-kDa protein. Northern blot hybridization using total RNA from several tissues of the rat provided evidence that the 14-kDa protein was expressed specifically in rat brain. Transfection of this cDNA into mammalian cells resulted in expression of the 14-kDa protein. The amino acid sequence predicted from the cDNA of the rat brain 14-kDa protein contained 137 amino acid residues. A hydropathy profile revealed a hydrophobic domain (amino acids 60-80) flanked by highly hydrophilic stretches on both sides. Whereas the N-terminal region of the 14-kDa protein contained four repeating motifs, EKTKEGV, the C-terminal domain was rich in glutamic acid and proline. A computer search of the amino acid sequence of the 14-kDa protein indicated no homology to any other protein reported so far.     

Arabidopsis Aux/IAA genes are involved in brassinosteroid-mediated growth responses in a manner dependent on organ type

We examined whether auxin/indole-3-acetic acid (Aux/IAA) proteins, which are key players in auxin-signal transduction, are involved in brassinosteroid (BR) responses. iaa7/axr2-1 and iaa17/axr3-3 mutants showed aberrant BR sensitivity and aberrant BR-induced gene expression in an organ-dependent manner. Two auxin inhibitors were tested in terms of BR responses. Yokonolide B inhibited BR responses, whereas p-chlorophenoxyisobutyric acid did not inhibit BR responses. DNA microarray analysis revealed that 108 genes were up-regulated, while only eight genes were down-regulated in iaa7. Among the genes that were up- or down-regulated in axr2, 22% were brassinolide-inducible genes, 20% were auxin-inducible genes, and the majority were sensitive neither to BR nor to auxin. An inhibitor of BR biosynthesis, brassinazole, inhibited auxin induction of the DR5-GUS gene, which consists of a synthetic auxin-response element, a minimum promoter, and a beta-glucuronidase. These results suggest that Aux/IAA proteins function in auxin- and BR-signaling pathways, and that IAA proteins function as the signaling components modulating BR sensitivity in a manner dependent on organ type.     

Aichi virus leader protein is involved in viral RNA replication and encapsidation

Aichi virus, a member of the family Picornaviridae, encodes a leader (L) protein of 170 amino acids (aa). The Aichi virus L protein exhibits no significant sequence homology to those of other picornaviruses. In this study, we investigated the function of the Aichi virus L protein in virus growth. In vitro translation and cleavage assays indicated that the L protein has no autocatalytic activity and is not involved in polyprotein cleavage. The L-VP0 junction was cleaved by 3C proteinase. Immunoblot analysis showed that the L protein is stably present in infected cells. Characterization of various L mutants derived from an infectious cDNA clone revealed that deletion of 93 aa of the center part (aa 43 to 135), 50 aa of the N-terminal part (aa 4 to 53), or 90 aa of the C-terminal part (aa 74 to 163) abolished viral RNA replication. A mutant (Delta114-163) in which 50 aa of the C-terminal part (aa 114 to 163) were deleted exhibited efficient RNA replication and translation abilities, but the virus yield was 4 log orders lower than that of the wild type. Sedimentation analysis of viral particles generated in mutant Delta114-163 RNA-transfected cells showed that the mutant has a severe defect in the formation of mature virions, but not in that of empty capsids. Thus, the data obtained in this study indicate that the Aichi virus L protein is involved in both viral RNA replication and encapsidation.