TRPA1 acts as a cold sensor in vitro and in vivo Karashima, Yuji; Talavera, Karel; Everaerts, Wouter ...
Proceedings of the National Academy of Sciences - PNAS,
01/2009, Volume:
106, Issue:
4
Journal Article
Peer reviewed
Open access
TRPA1 functions as an excitatory ionotropic receptor in sensory neurons. It was originally described as a noxious cold-activated channel, but its cold sensitivity has been disputed in later studies, ...and the contribution of TRPA1 to thermosensing is currently a matter of strong debate. Here, we provide several lines of evidence to establish that TRPA1 acts as a cold sensor in vitro and in vivo. First, we demonstrate that heterologously expressed TRPA1 is activated by cold in a Ca²⁺-independent and Ca²⁺ store-independent manner; temperature-dependent gating of TRPA1 is mechanistically analogous to that of other temperature-sensitive TRP channels, and it is preserved after treatment with the TRPA1 agonist mustard oil. Second, we identify and characterize a specific subset of cold-sensitive trigeminal ganglion neurons that is absent in TRPA1-deficient mice. Finally, cold plate and tail-flick experiments reveal TRPA1-dependent, cold-induced nociceptive behavior in mice. We conclude that TRPA1 acts as a major sensor for noxious cold.
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Astrocytes contribute to the formation and function of synapses and are found throughout the brain, where they show intracellular store-mediated Ca(2+) signals. Here, using a membrane-tethered, ...genetically encoded calcium indicator (Lck-GCaMP3), we report the serendipitous discovery of a new type of Ca(2+) signal in rat hippocampal astrocyte-neuron cocultures. We found that Ca(2+) fluxes mediated by transient receptor potential A1 (TRPA1) channels gave rise to frequent and highly localized 'spotty' Ca(2+) microdomains near the membrane that contributed appreciably to resting Ca(2+) in astrocytes. Mechanistic evaluations in brain slices showed that decreases in astrocyte resting Ca(2+) concentrations mediated by TRPA1 channels decreased interneuron inhibitory synapse efficacy by reducing GABA transport by GAT-3, thus elevating extracellular GABA. Our data show how a transmembrane Ca(2+) source (TRPA1) targets a transporter (GAT-3) in astrocytes to regulate inhibitory synapses.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The existence of non-canonical nicotinamide adenine diphosphate (NAD) 5'-end capped RNAs is now well established. Nevertheless, the biological function of this nucleotide metabolite cap remains ...elusive. Here, we show that the yeast Saccharomyces cerevisiae cytoplasmic 5'-end exoribonuclease Xrn1 is also a NAD cap decapping (deNADding) enzyme that releases intact NAD and subsequently degrades the RNA. The significance of Xrn1 deNADding is evident in a deNADding deficient Xrn1 mutant that predominantly still retains its 5'-monophosphate exonuclease activity. This mutant reveals Xrn1 deNADding is necessary for normal growth on non-fermenting sugar and is involved in modulating mitochondrial NAD-capped RNA levels and may influence intramitochondrial NAD levels. Our findings uncover a contribution of mitochondrial NAD-capped RNAs in overall NAD regulation with the deNADding activity of Xrn1 fulfilling a central role.
Sensorineural hearing loss is caused by the loss of sensory hair cells and neurons of the inner ear. Once lost, these cell types are not replaced. Two genes expressed in the developing inner ear are ...c-Myc and Sox2. We created immortalized multipotent otic progenitor (iMOP) cells, a fate-restricted cell type, by transient expression of C-MYC in SOX2-expressing otic progenitor cells. This activated the endogenous C-MYC and amplified existing SOX2-dependent transcripts to promote self-renewal. RNA-seq and ChIP-seq analyses revealed that C-MYC and SOX2 occupy over 85% of the same promoters. C-MYC and SOX2 target genes include cyclin-dependent kinases that regulate cell-cycle progression. iMOP cells continually divide but retain the ability to differentiate into functional hair cells and neurons. We propose that SOX2 and C-MYC regulate cell-cycle progression of these cells and that downregulation of C-MYC expression after growth factor withdrawal serves as a molecular switch for differentiation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
After endocytosis, transmembrane cargo reaches endosomes, where it encounters complexes dedicated to opposing functions: recycling and degradation. Microdomains containing endosomal sorting complexes ...required for transport (ESCRT)-0 component Hrs hepatocyte growth factor-regulated tyrosine kinase substrate (HGRS-1) in Caenorhabditis elegans mediate cargo degradation, concentrating ubiquitinated cargo and organizing the activities of ESCRT. At the same time, retromer associated sorting nexin one (SNX-1) and its binding partner, J-domain protein RME-8, sort cargo away from degradation, promoting cargo recycling to the Golgi. Thus, we hypothesized that there could be important regulatory interactions between retromer and ESCRT that balance degradative and recycling functions. Taking advantage of the naturally large endosomes of the C. elegans coelomocyte, we visualized complementary ESCRT-0 and RME-8/SNX-1 microdomains in vivo and assayed the ability of retromer and ESCRT microdomains to regulate one another. We found in snx-1(0) and rme-8(ts) mutants increased endosomal coverage and intensity of HGRS-1–labeled microdomains, as well as increased total levels of HGRS-1 bound to membranes. These effects are specific to SNX-1 and RME-8, as loss of other retromer components SNX-3 and vacuolar protein sorting-associated protein 35 (VPS-35) did not affect HGRS-1 microdomains. Additionally, knockdown of hgrs-1 had little to no effect on SNX-1 and RME-8microdomains, suggesting directionality to the interaction. Separation of the functionally distinct ESCRT-0 and SNX-1/RME-8 microdomains was also compromised in the absence of RME-8 and SNX-1, a phenomenonwe observed to be conserved, as depletion of Snx1 and Snx2 in HeLa cells also led to greater overlap of Rme-8 and Hrs on endosomes.
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•Many inner ear cell types express CHD4.•CHD4 activity at enhancers can regulate gene expression.•CHD4 suppresses alternative cell lineages to facilitate cell fate transitions.
Loss of spiral ...ganglion neurons (SGNs) in the cochlea causes hearing loss. Understanding the mechanisms of cell fate transition accelerates efforts that employ directed differentiation and lineage conversion to repopulate lost SGNs. Proposed strategies to regenerate SGNs rely on altering cell fate by activating transcriptional regulatory networks, but repressing networks for alternative cell lineages is also essential. Epigenomic changes during cell fate transitions suggest that CHD4 represses gene expression by altering the chromatin status. Despite limited direct investigations, human genetic studies implicate CHD4 function in the inner ear. The possibility of CHD4 in suppressing alternative cell fates to promote inner ear regeneration is discussed.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Transient receptor potential ankyrin 1 (TRPA1) is expressed by nociceptive neurons of the dorsal root ganglia (DRGs) and trigeminal ganglia, but its roles in cold and mechanotransduction are ...controversial. To determine the contribution of TRPA1 to cold and mechanotransduction in cutaneous primary afferent terminals, we used the ex vivo skin-nerve preparation from Trpa1(+/+), Trpa1(+/-), and Trpa1(-/-) adult mouse littermates. Cutaneous fibers from TRPA1-deficient mice showed no deficits in acute cold sensitivity, but they displayed striking deficits in mechanical response properties. C-fiber nociceptors from Trpa1(-/-) mice exhibited action potential firing rates 50% lower than those in wild-type C-fibers across a wide range of force intensities. Adelta-fiber mechanonociceptors also had reduced firing, but only at high intensity forces (>100 mN). Surprisingly, the firing rates of low-threshold Abeta and D-hair mechanoreceptive fibers were also altered. TRPA1 protein and mRNA expression was assessed in DRG neurons and cutaneous innervation by using Trpa1 in situ hybridization, an antibody for TRPA1, and an antibody for placental alkaline phosphatase (PLAP) in mice in which PLAP was substituted for Trpa1. DRG neurons of all sizes expressed Trpa1 mRNA or PLAP immunoreactivity. TRPA1 or PLAP immunolabeling was detected not only on many thin-caliber axons and intraepidermal endings but also on many large-caliber axons as well as lanceolate and Meissner endings. Epidermal and hair follicle keratinocytes also express TRPA1 message and protein. We propose that TRPA1 modulates mechanotransduction via a cell-autonomous mechanism in nociceptor terminals and possibly through a modulatory role in keratinocytes, which may interact with sensory terminals to modify their mechanical firing properties.
Microglia are critical in brain development and Alzheimer’s disease (AD) etiology. Down syndrome (DS) is the most common genetic developmental disorder and risk factor for AD. Surprisingly, little ...information is available on the impact of trisomy of human chromosome 21 (Hsa21) on microglial functions during DS brain development and in AD in DS. Using induced pluripotent stem cell (iPSC)-based organoid and chimeric mouse models, we report that DS microglia exhibit an enhanced synaptic pruning function, which alters neuronal synaptic functions. In response to human brain tissue-derived pathological tau, DS microglia undergo cellular senescence and exhibit elevated type-I-interferon signaling. Mechanistically, knockdown of Hsa21-encoded type I interferon receptors, IFNARs, rescues the DS microglial phenotypes both during brain development and in response to pathological tau. Our findings provide in vivo evidence that human microglia respond to pathological tau by exhibiting dystrophic phenotypes. Targeting IFNARs may improve DS microglial functions and prevent senescence.
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•DS microglia over-prune synapses in hiPSC-based organoid and chimeric mouse models•DS microglia show dystrophic and senescent phenotypes in response to pathological tau•IFN-I signaling is upregulated in DS microglia under pathophysiological conditions•Reducing IFNAR expression improves DS microglial functions and prevents senescence
With human iPSC-based brain organoids and mouse chimeras, Jin and colleagues demonstrate that upregulated type-I-interferon (IFN-I) signaling in Down syndrome microglia causes elevated synaptic pruning during development and accelerated senescence, rather than massive activation/inflammation, in response to pathological tau. These phenotypes can be rescued by inhibiting IFN-I receptors.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Mechanical stimuli generated by head movements and changes in sound pressure are detected by hair cells with amazing speed and sensitivity. The mechanosensitive organelle, the hair bundle, is a ...highly elaborated structure of actin-based stereocilia arranged in precise rows of increasing height. Extracellular linkages contribute to its cohesion and convey forces to mechanically gated channels. Channel opening is nearly instantaneous and is followed by a process of sensory adaptation that keeps the channels poised in their most sensitive range. This process is served by motors, scaffolds, and homeostatic mechanisms. The molecular constituents of this process are rapidly being elucidated, especially by the discovery of deafness genes and antibody targets.
Inner ear morphogenesis requires tightly regulated epigenetic and transcriptional control of gene expression. CHD7, an ATP-dependent chromodomain helicase DNA-binding protein, and SOX2, an ...SRY-related HMG box pioneer transcription factor, are known to contribute to vestibular and auditory system development, but their genetic interactions in the ear have not been explored. Here, we analyzed inner ear development and the transcriptional regulatory landscapes in mice with variable dosages of
and/or
. We show that combined haploinsufficiency for
and
results in reduced otic cell proliferation, severe malformations of semicircular canals, and shortened cochleae with ectopic hair cells. Examination of mice with conditional, inducible
loss by
reveals a critical period (~E9.5) of susceptibility in the inner ear to combined
and
loss. Data from genome-wide RNA-sequencing and CUT&Tag studies in the otocyst show that CHD7 regulates
expression and acts early in a gene regulatory network to control expression of key otic patterning genes, including
and
. CHD7 and SOX2 directly bind independently and cooperatively at transcription start sites and enhancers to regulate otic progenitor cell gene expression. Together, our findings reveal essential roles for
and
in early inner ear development and may be applicable for syndromic and other forms of hearing or balance disorders.
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