Capsaicin, a pungent ingredient of hot peppers, causes excitation of small sensory neurons, and thereby produces severe pain. A nonselective cation channel activated by capsaicin has been identified in sensory neurons and a cDNA encoding the channel has been cloned recently. However, an endogenous activator of the receptor has not yet been found. In this study, we show that several products of lipoxygenases directly activate the capsaicinactivated channel in isolated membrane patches of sensory neurons. Among them, 12-and 15-(S)-hydroperoxyeicosatetraenoic acids, 5-and 15-(S)-hydroxyeicosatetraenoic acids, and leukotriene B4 possessed the highest potency. The eicosanoids also activated the cloned capsaicin receptor (VR1) expressed in HEK cells. Prostaglandins and unsaturated fatty acids failed to activate the channel. These results suggest a novel signaling mechanism underlying the pain sensory transduction.
Capsaicin (CAP) excites small sensory neurons, causing pain, neurogenic inflammation, and other visceral reflexes. These effects have been proposed to be the result of CAP activation of a nonselective cation current. It is generally assumed that CAP binds to an extracellular domain of the membrane receptor. However, the exact binding site is not known because of the lipophilic nature of CAP. To determine whether the binding domain is extracellular or intracellular, we tested the effect of a synthetic watersoluble CAP analog, DA-5018⅐HCl, on current activation. CAP activated the 45 pS (at Ϫ60 mV) nonselective cation channel from either side of the membrane. However, DA-5018⅐HCl, which had a greater potency and efficacy than CAP, activated the channels only from the cytosolic side of the patch membrane in a capsazepine, a CAP receptor antagonist, reversible manner. When applied extracellularly, DA-5018⅐HCl did not, but CAP did, activate whole-cell currents in sensory neurons, as well as in oocytes expressing vanilloid receptor 1, a recently cloned CAP receptor. Hydrogen ions, reported as a possible endogenous activator of cation current, failed to elicit any current when acidic medium (pH 5.0-6.0) was applied intracellularly, indicating that H ϩ does not mediate the CAP effect. These results indicate that CAP and its analog bind to the cytosolic domain of the CAP receptor and suggest that an endogenous CAP-like substance other than H ϩ may be present in the cell.
Interleukin-2 (IL-2)-induced transcription of the J chain gene was used as a model for analyzing cytokine regulation during B cell development. To determine whether IL-2 signals are targeted to a J chain gene enhancer as well as to its promoter, the sequences flanking the J chain gene were first examined for DNase I hypersensitivity. Of six sites identified, two strong ones, 7.5 kb upstream of the J chain gene, were found to be associated with an enhancer that is active only during the antigen-driven stages of B cell development. Further analyses of the enhancer in the IL-2-responsive presecretor BCL1 cells showed that the enhancer is activated at this stage by an IL-2 signal that functions by opening the enhancer chromatin and stimulating STAT5 to bind to a STAT5 element critical for the enhancer induction. Moreover, after this early induction stage, the enhancer was shown to be constitutively open and active in terminally differentiated plasma cells.
TRPV1, a cloned capsaicin receptor, is a molecular sensor for detecting adverse stimuli and a key element for inflammatory nociception and represents biophysical properties of native channel. However, there seems to be a marked difference between TRPV1 and native capsaicin receptors in the pharmacological response profiles to vanilloids or acid. One plausible explanation for this overt discrepancy is the presence of regulatory proteins associated with TRPV1. Here, we identify Fas-associated factor 1 (FAF1) as a regulatory factor, which is coexpressed with and binds to TRPV1 in sensory neurons. When expressed heterologously, FAF1 reduces the responses of TRPV1 to capsaicin, acid, and heat, to the pharmacological level of native capsaicin receptor in sensory neurons. Furthermore, silencing FAF1 by RNA interference augments capsaicin-sensitive current in native sensory neurons. We therefore conclude that FAF1 forms an integral component of the vanilloid receptor complex and that it constitutively modulates the sensitivity of TRPV1 to various noxious stimuli in sensory neurons.
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