PubMed Health⌕ Search

Biomedical subjects

C Montell

Publications and source records attributed to C Montell.

At least 19 recordsLinked to original sources
PubMed2001

Regulation of the rhodopsin protein phosphatase, RDGC, through interaction with calmodulin.

Hundreds of G protein-coupled receptors (GPCRs) and at least six GPCR kinases have been identified, but the only GPCR phosphatase that has been definitively demonstrated is the rhodopsin phosphatase encoded by the rdgC locus of Drosophila. Mutations in rdgC result in defects in termination of the light response and cause severe retinal degeneration. In the current work, we demonstrate that RDGC binds to calmodulin, and a mutation in an IQ motif that eliminates the calmodulin/RDGC interaction prevents dephosphorylation of rhodopsin in vivo and disrupts termination of the photoresponse. Our data indicate that RDGC is a novel calmodulin-dependent protein phosphatase and raise the possibility that regulation of other GPCRs through dephosphorylation may be controlled by calmodulin-dependent protein phosphatases related to RDGC.

Amino Acid Sequence↗
PubMed2001

Regulation of melastatin, a TRP-related protein, through interaction with a cytoplasmic isoform.

The TRP (transient receptor potential) superfamily includes a group of subfamilies of channel-like proteins mediating a multitude of physiological signaling processes. The TRP-melastatin (TRPM) subfamily includes the putative tumor suppressor melastatin (MLSN) and is a poorly characterized group of TRP-related proteins. Here, we describe the identification and characterization of an additional TRPM protein TRPM4. We reveal that TRPM4 and MLSN each mediate Ca(2+) entry when expressed in HEK293 cells. Furthermore, we demonstrate that a short form of MLSN (MLSN-S) interacts directly with and suppresses the activity of full-length MLSN (MLSN-L). This suppression seems to result from the inhibition of translocation of MLSN-L to the plasma membrane. We propose that control of translocation through interaction between MLSN-S and MLSN-L represents a mode for regulating ion channel activity.

Amino Acid Sequence↗
PubMed2001

Physiology, phylogeny, and functions of the TRP superfamily of cation channels.

The transient receptor potential (TRP) protein superfamily consists of a diverse group of Ca(2+) permeable nonselective cation channels that bear structural similarities to Drosophila TRP. TRP-related proteins play important roles in nonexcitable cells, as demonstrated by the recent finding that a mammalian TRPC protein is expressed in endothelial cells and functions in vasorelaxation. However, an emerging theme is that many TRP-related proteins are expressed predominantly in the nervous system and function in sensory physiology. The TRP superfamily can be divided into six subfamilies, the first of which is composed of the "classical TRPs" (TRPC subfamily). These proteins all share the common features of three to four ankryin repeats, >/=30% amino acid homology over >/=750 amino acids, and a gating mechanism that operates through phospholipase C. Some classical TRPs may be store-operated channels (SOCs), which are activated by release of Ca(2+) from internal stores. The mammalian TRPC proteins are also expressed in the central nervous system, and several are highly enriched in the brain. One TRPC protein has been implicated in the pheromone response. The archetypal TRP, Drosophila TRP, is predominantly expressed in the visual system and is required for phototransduction. Many members of a second subfamily (TRPV) function in sensory physiology. These include VR1 and OSM-9, which respond to heat, osmolarity, odorants, and mechanical stimuli. A third subfamily, TRPN, includes proteins with many ankyrin repeats, one of which, NOMPC, participates in mechanotransduction. Among the members of a fourth subfamily, TRPM, is a putative tumor suppressor termed melastatin, and a bifunctional protein, TRP-PLIK, consisting of a TRPM channel fused to a protein kinase. PKD2 and mucolipidin are the founding members of the TRPP and TRPML subfamilies, respectively. Mutations in PKD2 are responsible for polycystic kidney disease, and mutations in mucolipidin result in a severe neurodegenerative disorder. Recent studies suggest that alterations in the activities of SOC and TRP channels may be at the heart of several additional neurodegenerative diseases. Thus, TRP channels may prove to be important new targets for drug discovery.

Animals↗
PubMed2001

Assessment of the role of the inositol 1,4,5-trisphosphate receptor in the activation of transient receptor potential channels and store-operated Ca2+ entry channels.

The mechanism for coupling between Ca(2+) stores and store-operated channels (SOCs) is an important but unresolved question. Although SOCs have not been molecularly identified, transient receptor potential (TRP) channels share a number of operational parameters with SOCs. The question of whether activation of SOCs and TRP channels is mediated by the inositol 1,4,5-trisphosphate receptor (InsP(3)R) was examined using the permeant InsP(3)R antagonist, 2-aminoethoxydiphenyl borate (2-APB) in both mammalian and invertebrate systems. In HEK293 cells stably transfected with human TRPC3 channels, the actions of 2-APB to block carbachol-induced InsP(3)R-mediated store release and carbachol-induced Sr(2+) entry through TRPC3 channels were both reversed at high agonist levels, suggesting InsP(3)Rs mediate TRPC3 activation. However, electroretinogram recordings of the light-induced current in Drosophila revealed that the TRP channel-mediated responses in wild-type as well as trp and trpl mutant flies were all inhibited by 2-APB. This action of 2-APB is likely InsP(3)R-independent since InsP(3)Rs are dispensable for the light response. We used triple InsP(3)R knockout DT40 chicken B-cells to further assess the role of InsP(3)Rs in SOC activation. (45)Ca(2+) flux analysis revealed that although DT40 wild-type cells retained normal InsP(3)Rs mediating 2-APB-sensitive Ca(2+) release, the DT40InsP(3)R-k/o cells were devoid of functional InsP(3)Rs. Using intact cells, all parameters of Ca(2+) store function and SOC activation were identical in DT40wt and DT40InsP(3)R-k/o cells. Moreover, in both cell lines SOC activation was completely blocked by 2-APB, and the kinetics of action of 2-APB on SOCs (time dependence and IC(50)) were identical. The results indicate that (a) the action of 2-APB on Ca(2+) entry is not mediated by the InsP(3)R and (b) the effects of 2-APB provide evidence for an important similarity in the function of invertebrate TRP channels, mammalian TRP channels, and mammalian store-operated channels.

Adenosine↗
PubMed2000

TRP and the PDZ protein, INAD, form the core complex required for retention of the signalplex in Drosophila photoreceptor cells.

The light response in Drosophila photoreceptor cells is mediated by a series of proteins that assemble into a macromolecular complex referred to as the signalplex. The central player in the signalplex is inactivation no afterpotential D (INAD), a protein consisting of a tandem array of five PDZ domains. At least seven proteins bind INAD, including the transient receptor potential (TRP) channel, which depends on INAD for localization to the phototransducing organelle, the rhabdomere. However, the determinants required for localization of INAD are not known. In this work, we showed that INAD was required for retention rather than targeting of TRP to the rhabdomeres. In addition, we demonstrated that TRP bound to INAD through the COOH terminus, and this interaction was required for localization of INAD. Other proteins that depend on INAD for localization, phospholipase C and protein kinase C, also mislocalized. However, elimination of any other member of the signalplex had no impact on the spatial distribution of INAD. A direct interaction between TRP and INAD did not appear to have a role in the photoresponse independent of localization of multiple signaling components. Rather, the primary function of the TRP/ INAD complex is to form the core unit required for localization of the signalplex to the rhabdomeres.

Age Factors↗
PubMed2000

TRPgamma, a drosophila TRP-related subunit, forms a regulated cation channel with TRPL.

TRP and TRPL are two light-sensitive cation channel subunits required for the Drosophila photoresponse; however, our understanding of the identities, subunit composition, and function of the light-responsive channels is incomplete. To explain the residual photoresponse that remains in the trp mutant, a third TRP-related subunit has previously been proposed to function with TRPL. Here, we identify such a subunit, TRPgamma. We show that TRPgamma is highly enriched in photoreceptor cells and preferentially heteromultimerizes with TRPL in vitro and in vivo. The N-terminal domain of TRPgamma dominantly suppressed the TRPL-dependent photoresponse, indicating that TRPgamma-TRPL heteromultimers contribute to the photoresponse. While TRPL and TRPgamma homomultimers are constitutively active, we demonstrate that TRPL-TRPgamma heteromultimers form a regulated phospholipase C- (PLC-) stimulated channel.

Amino Acid Sequence↗
PubMed1999

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.

Animals↗
PubMed1999

Termination of phototransduction requires binding of the NINAC myosin III and the PDZ protein INAD.

Many of the proteins that are critical for Drosophila phototransduction assemble into a signaling complex, signalplex, through association with the PDZ-domain protein INAD. Some of these proteins depend on INAD for proper subcellular localization to the phototransducing organelle, the rhabdomere, making it difficult to assess any physiological function of this signaling complex independent of localization. Here we demonstrated that INAD bound directly to the NINAC myosin III, yet the subcellular localization of NINAC was normal in inaD mutants. Nevertheless, the INAD binding site was sufficient to target a heterologous protein to the rhabdomeres. Disruption of the NINAC/INAD interaction delayed termination of the photoreceptor response. Thus one role of this signaling complex is in rapid deactivation of the photoresponse.

Animals↗
PubMed1999

Visual transduction in Drosophila.

The Drosophila phototransduction cascade has emerged as an attractive paradigm for understanding the molecular mechanisms underlying visual transduction, as well as other G protein-coupled signaling cascades that are activated and terminated with great rapidity. A large collection of mutants affecting the fly visual cascade have been isolated, and the nature and function of many of the affected gene products have been identified. Virtually all of the proteins, including those that were initially classified as novel, are highly related to vertebrate homologs. Recently, it has become apparent that most of the proteins central to Drosophila phototransduction are coupled into a supramolecular signaling complex, signalplex, through association with a PDZ-containing scaffold protein. The characterization of this complex has led to a re-evaluation of the mechanisms underlying the activation and deactivation of the phototransduction cascade.

Animals↗
PubMed1998

Requirement for the NINAC kinase/myosin for stable termination of the visual cascade.

Activation of the Drosophila photoresponse is a rapid process that results in plasma membrane Ca2+ and Na+ conductances. Ca2+ functions in negative feedback regulation of Drosophila vision including deactivation. Protein kinase C (PKC) binds directly to Ca2+ and is required for deactivation. However, the consequences of disrupting phosphorylation of any individual PKC substrate in the Drosophila retina have not been addressed. In the current work, we show that NINAC p174, which consists of a protein kinase domain joined to the head region of myosin heavy chain, is a phosphoprotein and is phosphorylated in vitro by PKC. Mutation of either of two PKC sites in the p174 tail resulted in an unusual defect in deactivation that had not been detected previously for other ninaC alleles or other loci. After cessation of the light stimulus, there appeared to be a transient reactivation of the visual cascade. This phenotype suggests that a mechanism exists to prevent reactivation of the visual cascade and that p174 participates in this process.

Animals↗
PubMed1998

Retinal targets for calmodulin include proteins implicated in synaptic transmission.

Ca2+ influxes regulate multiple events in photoreceptor cells including phototransduction and synaptic transmission. An important Ca2+ sensor in Drosophila vision appears to be calmodulin since a reduction in levels of retinal calmodulin causes defects in adaptation and termination of the photoresponse. These functions of calmodulin appear to be mediated, at least in part, by four previously identified calmodulin-binding proteins: the TRP and TRPL ion channels, NINAC and INAD. To identify additional calmodulin-binding proteins that may function in phototransduction and/or synaptic transmission, we conducted a screen for retinal calmodulin-binding proteins. We found eight additional calmodulin-binding proteins that were expressed in the Drosophila retina. These included six targets that were related to proteins implicated in synaptic transmission. Among these six were a homolog of the diacylglycerol-binding protein, UNC13, and a protein, CRAG, related to Rab3 GTPase exchange proteins. Two other calmodulin-binding proteins included Pollux, a protein with similarity to a portion of a yeast Rab GTPase activating protein, and Calossin, an enormous protein of unknown function conserved throughout animal phylogeny. Thus, it appears that calmodulin functions as a Ca2+ sensor for a broad diversity of retinal proteins, some of which are implicated in synaptic transmission.

Amino Acid Sequence↗
PubMed1998

Coordination of an array of signaling proteins through homo- and heteromeric interactions between PDZ domains and target proteins.

The rapid activation and feedback regulation of many G protein signaling cascades raises the possibility that the critical signaling proteins may be tightly coupled. Previous studies show that the PDZ domain containing protein INAD, which functions in Drosophila vision, coordinates a signaling complex by binding directly to the light-sensitive ion channel, TRP, and to phospholipase C (PLC). The INAD signaling complex also includes rhodopsin, protein kinase C (PKC), and calmodulin, though it is not known whether these proteins bind to INAD. In the current work, we show that rhodopsin, calmodulin, and PKC associate with the signaling complex by direct binding to INAD. We also found that a second ion channel, TRPL, bound to INAD. Thus, most of the proteins involved directly in phototransduction appear to bind to INAD. Furthermore, we found that INAD formed homopolymers and the homomultimerization occurred through two PDZ domains. Thus, we propose that the INAD supramolecular complex is a higher order signaling web consisting of an extended network of INAD molecules through which a G protein-coupled cascade is tethered.

Animals↗
PubMed1998

TRP trapped in fly signaling web.

In the Drosophila eye, photoactivation of rhodopsin leads to the opening of the light-sensitive cation influx channels TRP and TRPL. This response is extremely rapid and results in depolarization of the photoreceptor cells followed by Ca(2+)-mediated feedback regulation of the visual signaling cascade. The mechanisms that facilitate the rapid kinetics of activation and feedback regulation are poorly understood. However, the recent discovery that most of the proteins that function in fly phototransduction associate into a supramolecular complex permits a re-evaluation of the mechanisms underlying the activation and regulation of the cascade. The central player in the signaling complex is INAD, a protein with five protein-interaction motifs known as PDZ domains. The INAD complex does not appear to be a particle, but a massive signaling web composed of an INAD polymer with which some of the target proteins associate through complex multivalent interactions.

Animals↗
PubMed1997

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.

Animals↗
PubMed1997

Calmodulin regulation of light adaptation and store-operated dark current in Drosophila photoreceptors.

Phototransduction in Drosophila occurs through inositol lipid signaling that results in Ca2+ mobilization. In this system, we investigate the hitherto unknown physiological roles of calmodulin (CaM) in light adaptation and in regulation of the inward current that is brought about by depletion of cellular Ca2+ stores. To see the effects of a decreased Ca-CaM content in photoreceptor cells, we used several methods. Transgenic Drosophila P[ninaCDeltaB] flies, which have CaM-deficient photoreceptors, were studied. The peptide inhibitor M5, which binds to Ca-CaM and prevents its action, was applied. A Ca2+-free medium, which prevents Ca2+ influx and thereby diminishes the generation of Ca-CaM, was used. The decrease in the Ca-CaM level caused the following effects. (i) Fluorescence of Ca2+ indicator revealed an enhanced light-induced Ca2+ release from internal stores. (ii) Measurements of the light-induced current in P[ninaCDeltaB] cells showed a reduced light adaptation. (iii) Internal dialysis of M5 initially enhanced excitation and subsequently disrupted the light-induced current. (iv) An inward dark current appeared after depletion of the Ca2+ stores with ryanodine and caffeine. Importantly, application of Ca-CaM into the photoreceptor cells prevented all of the above effects. We propose that negative feedback of Ca-CaM on Ca2+ release from ryanodine-sensitive stores mediates light adaptation, is essential for light excitation, and keeps the store-operated inward current under a tight control.

Acclimatization↗