Craig Montell

Molecular characterization of the drosophila trp locus: A putative integral membrane protein required for phototransduction

Recent studies suggest that the fly uses the inositol lipid signaling system for visual excitation and that the Drosophila transient receptor potential (trp) mutation disrupts this process subsequent to the production of IP3. In this paper, we show that trp encodes a novel 1275 amino acid protein with eight putative transmembrane segments. Immunolocalization indicates that the trp protein is expressed predominantly in the rhabdomeric membranes of the photoreceptor cells.

The TRP Channels, a Remarkably Functional Family

TRP cation channels display an extraordinary assortment of selectivities and activation mechanisms, some of which represent previously unrecognized modes for regulating ion channels. Moreover, the biological roles of TRP channels appear to be equally diverse and range from roles in pain perception to male aggression.

A Unified Nomenclature for the Superfamily of TRP Cation Channels

The TRP superfamily includes a diversity of non-voltage-gated cation channels that vary significantly in their selectivity and mode of activation. Nevertheless, members of the TRP superfamily share significant sequence homology and predicted structural similarities. Currently, most of the genes and proteins that comprise the TRP superfamily have multiple names and, in at least one instance, two distinct genes belonging to separate subfamilies have the same name. Moreover, there are many cases in which highly related proteins that belong to the same subfamily have unrelated names. Therefore, to minimize confusion, we propose a unified nomenclature for the TRP superfamily.The current effort to unify the TRP nomenclature focuses on three subfamilies (TRPC, TRPV, and TRPM) that bear significant similarities to the founding member of this superfamily, Drosophila TRP, and which include highly related members in worms, flies, mice, and humans (Table 1)(Table 1). Members of the three subfamilies contain six transmembrane segments, a pore loop separating the final two transmembrane segments, and similarity in the lengths of the cytoplasmic and extracellular loops. In addition, the charged residues in the S4 segment that appear to contribute to the voltage sensor in voltage-gated ion channels are not conserved. The TRP-Canonical (TRPC) subfamily (formerly short-TRPs or STRPs) is comprised of those proteins that are the most highly related to Drosophila TRP. The TRPV subfamily (formerly OTRPC), is so named based on the original designation, Vanilloid Receptor 1 (VR1), for the first mammalian member of this subfamily (now TRPV1). The name for the TRPM subfamily (formerly long-TRPs or LTRPs) is derived from the first letter of Melastatin, the former name (now TRPM1) of the founding member of this third subfamily of TRP-related proteins. Based on amino acid homologies, the mammalian members of these three subfamilies can be subdivided into several groups each (Table 2Table 2 and Figure 1Figure 1) .Table 1Number of TRP Genes in Worms (C. elegans), Flies (Drosophila melanogaster), Mice, and HumansSubfamilyWormsFliesMiceHumansTRPC3376aaTRPV5255TRPM4188aTRPC2 is a pseudogene and is not counted.Table 2Nomenclature of the Mammalian TRP SuperfamilyNameGroupFormer NamesAccession NumbersTRPC11TRP1CAA61447, AAA93252TRPC1TRPC22TRP2X89067, AAD17195, AAD17196, AAG29950, AAG29951, AAD31453,TRPC2CAA06964TRPC33TRP3AAC51653TRPC3TRPC44TRP4CAA68125, BAA23599TRPC4TRPC54TRP5AAC13550, CAA06911, CAA06912TRPC5TRPC63TRP6NP_038866TRPC6TRPC73TRP7AAD42069, NP_065122TRPC7TRPV11VR1AAC53398OTRPC1TRPV21VRL-1AAD26363, AAD26364, BAA78478OTRPC2GRCTRPV3 (not assigned)TRPV42OTRPC4AAG17543, AAG16127, AAG28027, AAG28028, AAG28029,VR-OACCAC20703TRP12VRL-2TRPV53ECaC1CAB40138CaT2TRPV63CaT1AAD47636ECaC2CAC20416CaT-LCAC20417TRPM11MelastatinAAC13683, AAC80000TRPM22TRPC7BAA34700LTRPC2TRPM31KIAA1616AA038185LTRPC3TRPM43TRPM4H18835LTRPC4TRPM53MTR1AAF26288LTRPC5TRPM64Chak2AF350881TRPM74TRP-PLIKAAF73131Chak1LTRPC7TRPM82TRP-p8AC005538Indicated are the suggested gene and protein names, the groups within each subfamily, the former names, and accession numbers.Figure 1Phylogenetic Tree of the TRP SuperfamilyThe tree, which was adapted from Clapham et al., 2001 (Nat. Rev. Neurosci. 2, 387–396), was calculated using the neighbor-joining method and human, rat, and mouse sequences.View Large Image | View Hi-Res Image | Download PowerPoint SlideThe numbering system for the mammalian TRPC, TRPV, and TRPM proteins takes into account the order of their discovery and, in as many cases as possible, the number that has already been assigned to the genes and proteins (Table 2)(Table 2). In the case of the TRPV proteins, the numbering system is also based in part on the groupings of the TRPV proteins. New members of each subfamily will maintain the same root name and, with the exception of TRPV3, will be assigned the next number in the sequence. Currently, TRPV3 is unassigned to maintain the TRPV1/ TRPV2 and TRPV5/TRPV6 groupings and so that the former OTRPC4 could be renamed TRPV4. The next TRPV protein will be designated TRPV3.We hope this new nomenclature will add clarity to the field and simplify the naming of new members of the TRP superfamily. We recommend that accession numbers be used whenever it is necessary to unambiguously specify a given variant resulting from alternative mRNA splicing. Finally, this nomenclature has been approved by the HUGO Gene Nomenclature Committee and we recommend that this system be used in all future publications concerning TRPC, TRPV, and TRPM subfamily members.

Isolation of a Putative Phospholipase C Gene of Drosophila, norpA, and Its Role in Phototransduction

Severe norpA mutations in Drosophila eliminate the photoreceptor potential and render the fly completely blind. Recent biochemical analyses have shown that norpA mutants lack phospholipase C (PLC) activity in the eye. A combination of chromosomal walking and transposon-mediated mutagenesis was used to clone the norpA gene. This gene encodes a 7.5 kb RNA that is expressed in the adult head. In situ hybridizations of norpA cDNA to adult tissue sections show that this gene is expressed abundantly in the retina. The putative norpA protein is composed of 1095 amino acid residues and has extensive sequence similarity to a PLC amino acid sequence from bovine brain. We suggest that the norpA gene encodes a PLC expressed in the eye of Drosophila and that PLC is an essential component of the Drosophila phototransduction pathway.

p53 Mediates Cellular Dysfunction and Behavioral Abnormalities in Huntington’s Disease

We present evidence for a specific role of p53 in the mitochondria-associated cellular dysfunction and behavioral abnormalities of Huntingtons disease (HD). Mutant huntingtin (mHtt) with expanded polyglutamine (polyQ) binds to p53 and upregulates levels of nuclear p53 as well as p53 transcriptional activity in neuronal cultures. The augmentation is specific, as it occurs with mHtt but not mutant ataxin-1 with expanded polyQ. p53 levels are also increased in the brains of mHtt transgenic (mHtt-Tg) mice and HD patients. Perturbation of p53 by pifithrin-alpha, RNA interference, or genetic deletion prevents mitochondrial membrane depolarization and cytotoxicity in HD cells, as well as the decreased respiratory complex IV activity of mHtt-Tg mice. Genetic deletion of p53 suppresses neurodegeneration in mHtt-Tg flies and neurobehavioral abnormalities of mHtt-Tg mice. Our findings suggest that p53 links nuclear and mitochondrial pathologies characteristic of HD.

TRPM5 Is a Voltage-Modulated and Ca2+-Activated Monovalent Selective Cation Channel

The TRPM subfamily of mammalian TRP channels displays unusually diverse activation mechanisms and selectivities. One member of this subfamily, TRPM5, functions in taste receptor cells and has been reported to be activated through G protein-coupled receptors linked to phospholipase C. However, the specific mechanisms regulating TRPM5 have not been described. Here, we demonstrate that TRPM5 is a monovalent-specific cation channel with a 23 pS unitary conductance. TRPM5 does not display constitutive activity. Rather, it is activated by stimulation of a receptor pathway coupled to phospholipase C and by IP(3)-mediated Ca(2+) release. Gating of TRPM5 was dependent on a rise in Ca(2+) because it was fully activated by Ca(2+). Unlike any previously described mammalian TRP channel, TRPM5 displayed voltage modulation and rapid activation and deactivation kinetics upon receptor stimulation. The most closely related protein, the Ca(2+)-activated monovalent-selective cation channel TRPM4b, also showed voltage modulation, although with slower relaxation kinetics than TRPM5. Taken together, the data demonstrate that TRPM5 and TRPM4b represent the first examples of voltage-modulated, Ca(2+)-activated, monovalent cation channels (VCAMs). The voltage modulation and rapid kinetics provide TRPM5 with an excellent set of properties for participating in signaling in taste receptors and other excitable cells.

Activation of a TRPC3-dependent cation current through the neurotrophin BDNF.

Nonvoltage-gated cation currents, which are activated following stimulation of phospholipase C (PLC), appear to be major modes for Ca2+ and Na+ entry in mammalian cells. The TRPC channels may mediate some of these conductances since their expression in vitro leads to PLC-dependent cation influx. We found that the TRPC3 protein was highly enriched in neurons of the central nervous system (CNS). The temporal and spatial distribution of TRPC3 paralleled that of the neurotrophin receptor TrkB. Activation of TrkB by brain-derived nerve growth factor (BDNF) led to production of a PLC-dependent, nonselective cation conductance in pontine neurons. Evidence is provided that TRPC3 contributes to this current in vivo. Thus, activation of TrkB and PLC leads to a TRPC3-dependent cation influx in CNS neurons.

International Union of Pharmacology. XLIX. Nomenclature and Structure-Function Relationships of Transient Receptor Potential Channels

include a 25-amino acid (aa) motif in some subfamilies (the TRP domain) containing a TRP box (EWKFAR) just C-terminal to S6. The TRP domain and box, as well as slight variations of these motifs, are present in all TRPC and TRPM channel genes, but not in other TRP channels. The N-terminal cytoplasmic domains of TRPC, TRPV, and TRPA channels contain ankyrin repeats, whereas those of the TRPC and TRPM channels contain proline-rich sequences in the region just C-terminal portion of the TRP domain, referred to as TRP box 2 (Montell, 2005). At present, no features, other than overall 6TM architecture/homology and cationic permeability, define the TRP family. Thus the definition of TRP channels will evolve as functions and structures are clarified. Genes for the TRP ion channel subunits were first defined in the Drosophila visual system. In the trp mutant, the light response (receptor potential) decays dur

Coassembly of TRP and TRPL Produces a Distinct Store-Operated Conductance

The Drosophila retinal-specific protein, TRP (transient receptor potential), is the founding member of a family of store-operated channels (SOCs) conserved from C. elegans to humans. In vitro studies indicate that TRP is a SOC, but that the related retinal protein, TRPL, is constitutively active. In the current work, we report that coexpression of TRP and TRPL leads to a store-operated, outwardly rectifying current distinct from that owing to either TRP or TRPL alone. TRP and TRPL interact directly, indicating that the TRP-TRPL-dependent current is mediated by heteromultimeric association between the two subunits. We propose that the light-activated current in photoreceptor cells is produced by a combination of TRP homo- and TRP-TRPL heteromultimers.

Requirement for the PDZ Domain Protein, INAD, for Localization of the TRP Store-Operated Channel to a Signaling Complex

In Drosophila, the store-operated Ca2+ channel, TRP, is required in photoreceptor cells for a sustained response to light. Here, we show that TRP forms a complex with phospholipase C-beta (NORPA), rhodopsin (RH1), calmodulin, and the PDZ domain containing protein INAD. Proteins with PDZ domains have previously been shown to cluster ion channels in vitro. We show that in InaD mutant flies, TRP is no longer spatially restricted to its normal subcellular compartment, the rhabdomere. These results provide evidence that a PDZ domain protein is required, in vivo, for anchoring of an ion channel to a signaling complex. Furthermore, disruption of this interaction results in retinal degeneration. We propose that the TRP channel is linked to NORPA and RH1 to facilitate feedback regulation of these upstream signaling molecules.