InteractiveFly: GeneBrief

Transient receptor potential cation channel γ: Biological Overview | References

Gene name - Transient receptor potential cation channel γ

Synonyms - TRPγ

Cytological map position -

Function - Ca++ channel protein

Keywords - Ca++ channel protein activated by polyunsaturated fatty acids generated in a phospholipase C- and phospholipase A2-dependent manner, motor coordination, olfactory sensitivity, Malpighian tubule fluid transport, photoresponse

Symbol - Trpγ

FlyBase ID: FBgn0032593

Genetic map position - chr2L:16,663,454-16,676,701

Classification - Transient-receptor-potential calcium channel protein

Cellular location - surface transmembrane

NCBI link: EntrezGene
Trpgamma orthologs: Biolitmine
Recent literature
Riehle, M., Tsvetkov, D., Gohlke, B. O., Preissner, R., Harteneck, C., Gollasch, M. and Nurnberg, B. (2018). Molecular basis for the sensitivity of TRP channels to polyunsaturated fatty acids. Naunyn Schmiedebergs Arch Pharmacol. Pubmed ID: 29736621
Transient receptor potential (TRP) channels represent a superfamily of unselective cation channels that are subdivided into seven subfamilies based on their sequence homology and differences in gating and functional properties. Little is known about the molecular mechanisms of TRP channel regulation, particularly of the "canonical" TRP (TRPC) subfamily and their activation by polyunsaturated fatty acids (PUFAs). This study analyzed the structure-function relationship of Drosophila fruit fly TRPC channels. The primary aim was to uncover the molecular basis of PUFA sensitivity of Drosophila TRP-like (TRPL) and TRPgamma channels. Amino acid (aa) sequence alignment of the three Drosophila TRPC channels revealed 50 aa residues highly conserved in PUFA-sensitive TRPL and TRPgamma channels but not in the PUFA-insensitive TRP channel. Substitution of respective aa in TRPL by corresponding aa of TRP identified 18 residues that are necessary for PUFA-mediated activation of TRPL. Most aa positions are located within a stretch comprising transmembrane domains S2-S4, whereas six aa positions have been assigned to the proximal cytosolic C-terminus. Interestingly, residues I465 and S471 are required for activation by 5,8,11,14-eicosatetraynoic acid (ETYA) but not 5,8,11-eicosatriynoic acid (ETI). As proof of concept, a PUFA-sensitive TRP channel was generated by exchanging the corresponding aa from TRPL to TRP. This study demonstrates a specific aa pattern in the transmembrane domains S2-S4 and the proximal C-terminus essential for TRP channel activation by PUFAs.
Dhakal, S., Ren, Q., Liu, J., Akitake, B., Tekin, I., Montell, C. and Lee, Y. (2022). Drosophila TRPg is required in neuroendocrine cells for post-ingestive food selection. Elife 11. PubMed ID: 35416769
The mechanism through which the brain senses the metabolic state, enabling an animal to regulate food consumption, and discriminate between nutritional and non-nutritional foods is a fundamental question. Flies choose the sweeter non-nutritive sugar, L-glucose, over the nutritive D-glucose if they are not starved. However, under starvation conditions, they switch their preference to D-glucose, and this occurs independent of peripheral taste neurons. This study found that eliminating the TRPgamma channel impairs the ability of starved flies to choose D-glucose. This food selection depends on trpgamma expression in neurosecretory cells in the brain that express Diuretic hormone 44 (DH44). Loss of trpgamma increases feeding, alters the physiology of the crop, which is the fly stomach equivalent, and decreases intracellular sugars and glycogen levels. Moreover, survival of starved trpgamma flies is reduced. Expression of trpgamma in DH44 neurons reverses these deficits. These results highlight roles for TRPgamma in coordinating feeding with the metabolic state through expression in DH44 neuroendocrine cells.
Bajar, B. T., Phi, N. T., Isaacman-Beck, J., Reichl, J., Randhawa, H. and Akin, O. (2022). A discrete neuronal population coordinates brain-wide developmental activity. Nature 602(7898): 639-646. PubMed ID: 35140397
In vertebrates, stimulus-independent activity accompanies neural circuit maturation throughout the developing brain. The recent discovery of similar activity in the developing Drosophila central nervous system suggests that developmental activity is fundamental to the assembly of complex brains. How such activity is coordinated across disparate brain regions to influence synaptic development at the level of defined cell types is not well understood. This study shows that neurons expressing the cation channel transient receptor potential gamma (Trpγ) relay and pattern developmental activity throughout the Drosophila brain. In trpγ mutants, activity is attenuated globally, and both patterns of activity and synapse structure are altered in a cell-type-specific manner. Less than 2% of the neurons in the brain express Trpγ. These neurons arborize throughout the brain, and silencing or activating them leads to loss or gain of brain-wide activity. Together, these results indicate that this small population of neurons coordinates brain-wide developmental activity. It is proposed that stereotyped patterns of developmental activity are driven by a discrete, genetically specified network to instruct neural circuit assembly at the level of individual cells and synapses. This work establishes the fly brain as an experimentally tractable system for studying how activity contributes to synapse and circuit formation.
Palacios-Munoz, A., de Paula Moreira, D., Silva, V., Garcia, I. E., Aboitiz, F., Zarrei, M., Campos, G., Rennie, O., Howe, J. L., Anagnostou, E., Ambrozewic, P., Scherer, S. W., Passos-Bueno, M. R. and Ewer, J. (2022). Mutations in trpgamma, the homologue of TRPC6 autism candidate gene, causes autism-like behavioral deficits in Drosophila. Mol Psychiatry. PubMed ID: 35501408
Autism Spectrum Disorder (ASD) is characterized by impaired social communication, restricted interests, and repetitive and stereotyped behaviors. The TRPC6 (transient receptor potential channel 6) represents an ASD candidate gene under an oligogenic/multifactorial model based on the initial description and cellular characterization of an individual with ASD bearing a de novo heterozygous mutation disrupting TRPC6, together with the enrichment of disruptive TRPC6 variants in ASD cases as compared to controls. This study performed a clinical re-evaluation of the initial non-verbal patient, and also present eight newly reported individuals ascertained for ASD and bearing predicted loss-of-function mutations in TRPC6. In order to understand the consequences of mutations in TRPC6 on nervous system function, the fruit fly, Drosophila melanogaster, was used to show that null mutations in transient receptor gamma (trpγ the fly gene most similar to TRPC6), cause a number of behavioral defects that mirror features seen in ASD patients, including deficits in social interactions (based on courtship behavior), impaired sleep homeostasis (without affecting the circadian control of sleep), hyperactivity in both young and old flies, and defects in learning and memory. Some defects, most notably in sleep, differed in severity between males and females and became normal with age. Interestingly, hyperforin, a TRPC6 agonist and the primary active component of the St. John's wort antidepressant, attenuated many of the deficits expressed by trpγ mutant flies. In summary, these results provide further evidence that the TRPC6 gene is a risk factor for ASD. In addition, they show that the behavioral defects caused by mutations in TRPC6 can be modeled in Drosophila, thereby establishing a paradigm to examine the impact of mutations in other candidate genes.
Tian, Q., Wang, P., Xie, C., Pang, P., Zhang, Y., Gao, Y., Cao, Z., Wu, Y., Li, W., Zhu, M. X., Li, D. and Yao, J. (2022). Identification of an arthropod molecular target for plant-derived natural repellents. Proc Natl Acad Sci U S A 119(18): e2118152119. PubMed ID: 35452331
Arthropods maintain ecosystem balance while also contributing to the spread of disease. Plant-derived natural repellents represent an ecological method of pest control, but their direct molecular targets in arthropods remain to be further elucidated. Occupying a critical phylogenetic niche in arthropod evolution, scorpions retain an ancestral genetic profile. In this study, using a behavior-guided screening of the Mesobuthus martensii genome, a scorpion transient receptor potential (sTRP1) channel was identified that senses Cymbopogon-derived natural repellents, while remaining insensitive to the synthetic chemical pesticide DEET. Scrutinizing orthologs of sTRP1 in Drosophila melanogaster, we further demonstrated dTRPγ ion channel as a chemosensory receptor of natural repellents to mediate avoidance behavior. This study sheds light on arthropod molecular targets of natural repellents, exemplifying the arthropod–plant adaptation. It should also help the rational design of insect control strategy and in conserving biodiversity.

Motor coordination is broadly divided into gross and fine motor control, both of which depend on proprioceptive organs. However, the channels that function specifically in fine motor control are unknown. This study shows that mutations in trpγ disrupt fine motor control while leaving gross motor proficiency intact. The mutants are unable to coordinate precise leg movements during walking, and are ineffective in traversing large gaps due to an inability in making subtle postural adaptations that are requisite for this task. TRPγ is expressed in proprioceptive organs, and is required in both neurons and glia for gap crossing. TRPγ was expressed in vitro, and its activity was found to be promoted by membrane stretch. A mutation eliminating the Na+/Ca+ exchanger suppresses the gap-crossing phenotype of trpγ flies. These findings indicate that TRPγ contributes to fine motor control through mechanical activation in proprioceptive organs, thereby promoting Ca+ influx, which is required for function (Akitake, 2015).

Even the most basic tasks, such as acquiring food, locating safe places to rest, avoiding and defending against enemies, and mating requires motile animals to navigate through their environment by moving multiple body parts in a highly coordinated manner. To move fluidly, both vertebrate and invertebrate animals employ complex mechanosensory organs that are designed to gather and interpret feedback information about their movement in real time through an array of specialized receptors and neural networks. These proprioceptive sensory systems provide animals with continuously updated maps of their body positions that are critical for balance and locomotion (Akitake, 2015).

Proprioception is mediated at the cellular level, by stretch-sensitive cells located in muscles, ligaments and joints, which are activated by mechanical forces. In humans, damage to proprioceptive afferents results in a variety of movement disorders such as spasticity, impaired load sensitivity, and altered gait. Proprioceptive dysfunction is also a clinical feature of diseases that affect the nervous system such as Parkinson's disease (Akitake, 2015).

The worm, C. elegans, and the fruit fly, Drosophila melanogaster, have served as animal models for characterizing proprioception. Both of these organisms display highly stereotypic locomotion, which has facilitated the identification of neurons and ion channels that function in proprioception. In flies, proprioceptive neurons are located in specialized sensory structures-mechanosensory bristles, campaniform sensilla, and chordotonal organs. Several invertebrate members of the transient receptor potential (TRP) family of cation channels localize to proprioceptive cells and contribute to sensing bodily movements during locomotion. These include the C. elegans and Drosophila TRPN channels, TRP-4 and NOMPC, respectively, which are required for worms and fly larvae to make gross postural changes during locomotion. Most NompC mutant animals die during the pupal stage. The few mutant animals that survive to adulthood exhibit severe locomotion defects and uncoordinated movement of body parts, indicative of defects in gross motor control. Mutations disrupting the Drosophila TRPV channels, Inactive (Iav) and Nanchung (Nan) also result in severe locomotor defects (Akitake, 2015).

A key question is whether ion channels exist that specifically function in fine motor control. In flies, a defect in fine motor control would not eliminate behaviours that rely principally on gross movements of the body and appendages, such as negative geotaxis, or crossing small gaps. However, loss of fine motor control would be expected to impair performance when the flies are faced with highly challenging tasks, such as traversing wide gaps, which rely on coordinating a repertoire of fine movements, including subtle changes in body angles and leg positions (Akitake, 2015).

The Drosophila genome encodes 13 TRPs, 12 of which have been subjected to genetic analyses. The recurring theme is that these channels are essential for sensory physiology. However, the function of one Drosophila TRP channel, TRPγ, is not known. TRPγ is a TRPC channel (Xu, 2000), and is most related to the founding TRP channel. This study demonstrates that TRPγ is localized to neurons and glia that comprise the femoral chordotonal organs (FCOs). trpγ null mutant flies were generated and were found to be distinct from the nan and iav mutants in that they displayed much greater levels of negative geotaxis and were proficient in crossing small gaps. However, once the gaps became challenging but were still surmountable for most wild-type flies, the trpγ mutants were unable to make the fine postural adaptations required for negotiating these gaps. Thus, this phenotype sharply contrasted with the loss of other TRP channels that impact on proprioception, as TRPγ was uniquely required to promote this highly coordinated motor control. These data demonstrate that fine motor control is not mediated exclusively through the same repertoire of cation channels that function in gross motor control (Akitake, 2015).

In humans, motor control is categorized into two major types: gross motor skills (required for large body movements such as sitting upright or waving an arm), and fine motor skills, necessary for small precise movements such as picking up and manipulating objects. In Drosophila, gross motor coordination is essential for the large rhythmic movements of the body and limbs during general locomotion (walking), while fine motor coordination is critical for making small changes in the angles and positions of the body and appendages to complete difficult tasks, such as righting or navigating gaps in a terrain (Akitake, 2015).

Three Drosophila TRP channels are expressed in proprioceptive organs (Iav, Nan and NOMPC) and elimination of any of these proteins have severe effects on locomotion and gravity sensation. Owing to the major locomotor deficits resulting from the loss of any of these channels, fine motor control is also profoundly affected (Akitake, 2015).

Before the current study, it was unclear whether there existed cation channels that contribute exclusively to highly coordinated movements of the body and appendages. In principle, it was possible that the two types of motor coordination depended on the same repertoire of channels (for example, Iav and Nan), and that null mutations would strongly impair all types of coordinated movements, while hypomorphic mutations would affect fine motor control only (Akitake, 2015).

This study found that TRPγ was required exclusively for fine motor behaviours. In high-frame rate video analysis, both wild-type and trpγ flies traversed the catwalk at a relatively fixed maximum speed, although the trpγ mutants walked more slowly and displayed decreased precision in their leg placements. However, the trpγ mutants used consistently shortened steps, which differed from the abnormally long and highly variable step lengths exhibited by nan mutants11 (Akitake, 2015).

The impairments in fine motor control exhibited by trpγ flies compromised their ability to cross challenging gap sizes. While the mutant animals negotiated gaps of up to 3.0 mm as well as wild type, they were ineffective in traversing larger gaps. This defect was not due to smaller flies, since the lengths of the trpγ bodies were similar to wild-type animals. It is proposed that the impairment in gap-crossing arises from the inability of the mutants to precisely sense their body position and make the fine postural adjustments required to complete the task. Indeed, the trpγ flies were unable to increase their body angles towards the horizontal position, even as they made successive leg-over-head sweeps. Consequently, they could not fully extend the reach of their front legs to bridge the gap. In sharp contrast, nan and iav mutants were not able to effectively cross even short gaps that virtually all trpγ and wild-type flies were able to negotiate (Akitake, 2015).

The majority of work on Drosophila proprioceptive organs has focused on the contribution of the mechanosensory neurons to motor control. Unexpectedly, this study found that TRPγ was expressed and functioned in both neurons and in glial support cells, called scolopale cells. However, the requirement for TRPγ in neurons appeared to be more significant. RNAi knockdown of trpγ in neurons induced a gap-crossing deficit nearly as severe as the null mutations, while RNAi knockdown of trpγ in scolopale cells, caused a more modest effect. Moreover, the gap-crossing impairment exhibited by trpγ was rescued to a greater extent after re-introducing the wild-type transgene in neurons than in scolopale cells. When the effects of the trpγ mutation on the leg motor circuit were assayed, using EMGs, the deficit in sensitivity was rescued only after expressing the wild-type transgene in neurons. Nevertheless, TRPγ has a dual role in both neurons and scolopale cells, and this is an additional feature that distinguishes TRPγ from TRPs that function in gross motor control (Akitake, 2015).

In neurons, the spatial distribution of TRPγ is different from that of Iav, Nan and NOMPC, consistent with the distinct roles of these channels in promoting fine and gross motor control, respectively. In contrast to the cilia-restricted localizations of Iav, Nan, and NOMPC, detected TRPγ was detected throughout the neuronal cell bodies and dendrites (Akitake, 2015).

It is proposed that TRPγ functions in chordotonal neurons to sense joint movements needed for fine motor control. In support of this proposal, TRPγ was activated directly by membrane stretch in vitro, and expressed in the dendrites. Furthermore, the TRPγ-dependent Ca2+-influx contributes to function, since the severity of the behavioural phenotype was suppressed by eliminating the Na+/Ca2+-exchanger, CalX (Akitake, 2015).

An additional question concerns the potential role of TRPγ in the scolopale cells. This study found that the extensive vacuole network of the mutant scolopale cells in the FCO was reduced in size compared with the wild-type. Various mechanosensors contribute to maintaining vacuolar structures in cells and growing evidence suggests that TRP channels play critical roles in regulating cell size and shape. This raises the possibility that TRPγ plays a similar role in these support cells, which in turn helps maintain the structural stability of the mechanosensory organs (Akitake, 2015).

In summary, this study employed high-frame rate video microscopy of fly locomotive behaviours to identify a requirement for a mechanosensitive TRP channel, TRPγ, for fine motor control. The demonstration that a Drosophila channel functions specifically to promote precise body movements raises the question as to whether there exist mechanosensitive Ca2+-permeable channels in mammals that are uniquely required for fine motor control (Akitake, 2015).

Mutants in Drosophila TRPC channels reduce olfactory sensitivity to carbon dioxide

Members of the canonical Transient Receptor Potential (TRPC) class of cationic channels function downstream of Gαq and PLCβ in Drosophila photoreceptors for transducing visual stimuli. Gαq has recently been implicated in olfactory sensing of carbon dioxide (CO2) and other odorants. This study investigated the role of PLCβ and TRPC channels for sensing CO2 in Drosophila. Through behavioral assays it was demonstrated that Drosophila mutants for plc21c, trp and trpl have a reduced sensitivity for CO2. Immuno-histochemical staining for TRP, TRPL and TRPγ indicates that all three channels are expressed in Drosophila antennae including the sensory neurons that express CO2 receptors. Electrophysiological recordings obtained from the antennae of protein null alleles of TRP (trp343) and TRPL (trpl302), showed that the sensory response to multiple concentrations of CO2 was reduced. However, trpl302; trp343 double mutants still have a residual response to CO2. Down-regulation of TRPC channels specifically in CO2 sensing olfactory neurons reduced the response to CO2 and this reduction was obtained even upon down-regulation of the TRPCs in adult olfactory sensory neurons. Thus the reduced response to CO2 obtained from the antennae of TRPC RNAi strains is not due to a developmental defect. These observations show that reduction in TRPC channel function significantly reduces the sensitivity of the olfactory response to CO2 concentrations of 5% or less in adult Drosophila. It is possible that the CO2 receptors Gr63a and Gr21a activate the TRPC channels through Gαq and PLC21C (Badsha, 2012).

The role for TRPC channels in maintaining the high sensitivity of CO2 detection is important in multiple contexts. Detection of low concentrations of CO2 (5% or less) shares several similarities with odor detection. Receptors for low concentrations of CO2, despite belonging to the gustatory class of insect chemosensory receptors, are located within olfactory sensillae on the third antennal segment. Moreover, mutants in dgq, the gene that encodes the α subunit of the heterotrimeric G-protein Gαq, reduce the physiological response recorded from sensory neurons in both cases. This study shows that mutants of the ubiquitously expressed allele of PLCβ, plc21C (Shortridge, 1991) reduce the response to CO2 similar to the observation for odors (Kain, 2008) unlike mutants of norpA allele which is expressed strongly in the eyes and is required for phototransduction but not for CO2 sensing. In olfactory sensory neurons it has been proposed that the physiological response to odorants is a combination of ionotropic and metabotropic receptor signaling. The olfactory receptor and olfactory co-receptor (Or/Orco) complex forms an odor-activated ion channel in heterologous systems and is therefore thought to be an ionotropic component, while the olfactory receptor coupling to a G-protein, like Gαq, could initiate the metabotropic component through as yet un-determined ion channels. Unlike olfactory sensory neurons, ab1C, the CO2 sensing neurons do not express the olfactory receptor and olfactory co-receptor (Or/Orco) complex. Therefore, in these neurons it is possible that the ionotropic component is absent. The current data suggest that TRPCs, which are known to function downstream of Gq/Plcβ signaling may contribute to metabotropic signaling in ab1C neurons but the data does not allow this to be stated conclusively. However it is evident that the TRPC channels are required for the normal functioning of CO2 sensing ab1C neurons in adult Drosophila. The presence of a basal response in individual knock outs and knock downs of trp, trpl and trpγ and double null mutants of trp and trpl as compared to the complete lack of response in Gr63a null flies suggests that the CO2 sensing ab1C neurons are not solely dependent on the TRPC channels for function. While it is formally possible that the remaining response in trpl302;trp343 double nulls is due to trpγ, this idea is not favored primarily because, the response of double mutant nulls was no worse than that of single mutants. The triple mutant combination of trpl302;trp343 with the trpγ RNAi line was poorly viable and hence could not be tested directly (Badsha, 2012).

The consequences of this finding are relevant for Drosophila behavior. Unlike other insect species like moths and mosquitoes, Drosophila are innately repelled by low concentrations of CO2 presumably because it is an indicator of stress due to a potential threat to naïve flies. However, in conditions where CO2 is present along with food odorants this repulsion needs to be suppressed. The data suggest that TRPC channels are a component of this dual sensitivity. Repression of Gq/PLCβ signaling and/or TRPCs through mechanisms yet to be identified might reduce the sensitivity to CO2 and alter the behavior from repulsion to attraction. Interestingly, food odors that can reduce CO2 responses from ab1C neurons have been identified. Whether these odorants act through repression of TRPCs needs to be determined. Thus it appears that the three Drosophila TRPC channels TRP, TRPL and TRPγ can act as amplifiers of the signal downstream of a channel yet to be identified while playing redundant roles in this amplification process. The requirement for redundancy might stem from an evolutionarily conserved need to escape stress and or the necessity to find food (Badsha, 2012).

Receptor-induced activation of Drosophila TRP γ by polyunsaturated fatty acids

Cellular calcium homeostasis is regulated by hormones and neurotransmitters, resulting in the activation of a variety of proteins, in particular, channel proteins of the plasma membrane and of intracellular compartments. Such channels are, for example, TRP channels of the TRPC protein family that are activated by various mediators from receptor-stimulated signaling cascades. In Drosophila, two TRPC channels, TRP and TRPL, are involved in phototransduction. In addition, a third Drosophila TRPC channel, TRPγ, has been identified and described as an auxiliary subunit of TRPL. Beyond it, data show that heterologously expressed TRPγ formed a receptor-activated, outwardly rectifying cation channel independent from TRPL co-expression. Analysis of the activation mechanism revealed that TRPγ is activated by various polyunsaturated fatty acids generated in a phospholipase C- and phospholipase A(2)-dependent manner. The most potent activator of TRPγ, the stable analogue of arachidonic acid, 5,8,11,14-eicosatetraynoic acid, induced currents in single channel recordings. This study shows that upon heterologous expression TRPγ forms a homomeric channel complex that is activated by polyunsaturated fatty acids as mediators of receptor-dependent signaling pathways. Reverse transcription PCR analysis showed that TRPγ is expressed in Drosophila heads and bodies. Its body-wide expression pattern and its activation mechanism suggest that TRPγ forms a fly cation channel responsible for the regulation of intracellular calcium in a variety of hormonal signaling cascades (Jors, 2006).

TRPγ, as a member of the TRPC channel family, has been discussed as participating in Drosophila phototransduction. In the first report describing TRPγ, its expression was shown to specifically occur in Drosophila eyes and the head, but not in the body (Xu, 2000), suggesting a possible sensory function of TRPγ. In contrast, a second report showed that the transcription of TRPγ mRNA not only occurs in the fruit fly's head but also in the cells of the Malpighian tubules (MacPherson, 2005). Similarly, the current RT-PCR data support the notion that TRPγ expression is not restricted to Drosophila head. The fact that there are only three TRPC members (TRP, TRPL, TRPγ) in the genome of Drosophila and the expression of TRPγ in Drosophila heads and bodies make it likely that TRPγ-mediated Ca2+ influx is integrated in many receptor-mediated signaling pathways as shown for the mammalian TRPC channel proteins. Indeed, TRPγ, when heterologously expressed in HEK293 cells, can be regulated by hormonal activation of the cells via endogenous muscarinic GPCR, because both Ca2+-imaging and whole-cell patch clamp experiments showed an increased influx into TRPγ-expressing cells (Jors, 2006).

The signaling pathway leading to TRPγ activation was clarified with the help of pharmacological tools. The data in this study revealed that TRPγ, like TRPL, is activated by various polyunsaturated fatty acids generated in a phospholipase C- and phospholipase A2-dependent manner. Whereas the expression of a phospholipase C isoenzyme in Drosophila has been known since 1988, a gene coding for a phospholipase A2 isoenzyme has been found only recently (Chiang, 2004). Its gene product, called Radish, has been shown to be expressed in a specific subset of neurons involved in synaptic transmission necessary for olfactory memory. However, due to the lack of histological localization of TRPγ, it remains unclear whether TRPγ and Radish are involved in olfactory memory (Jors, 2006).

The PLA2 inhibitors used in this study have been shown to be specific for different PLA2 isoenzymes. N-(p-Amylcinnamoyl)anthranilic acid has been used as a common inhibitor, whereas p-bromphenacyl bromide, bromoenol lactone, and arachidonyltrifluoromethyl ketone have been described to be specific for the secretory, inducible, and cytosolic PLA2, respectively. Differences in inhibiting the carbachol-induced and TRPγ- or TRPL-mediated Ca2+ influx by these PLA2 inhibitors are, therefore, likely due to their different efficacies in inhibiting the endogenously expressed PLA2 in HEK293 cells (Jors, 2006).

The concentration-response relations of PUFAs show that TRPγ and TRPL are activated by polyunsaturated fatty acids with comparable efficacies, whereas palmitoleic acid, a monounsaturated fatty acid, is ineffective. Both channel proteins are stimulated not only by naturally occurring polyunsaturated fatty acids, ETYA, a synthetic PUFA analogue, also induced TRPγ currents. After the application of ETYA, current-voltage relationships were obtained similar to those measured under carbachol (Jors, 2006).

TRPγ in HEK293 cells was constitutively active, with a current-voltage relationship reversing around 0 mV and a permeability for cations PNa:PCs:PNMDG = 1:1:0.15, characterizing TRPγ as a non-selective cation channel. This is in good agreement with earlier data of Xu (2000). The amplitudes of inward and outward currents increased within 30 s after the application of carbachol, indicating that TRPγ is activated by mediators generated by intracellular signaling cascades. The ETYA-induced currents showed a current-voltage relationship and single channel conductance that is well comparable with that of the spontaneous currents (Jors, 2006).

The data on TRPγ obtained in this study, together with those for TRPL and TRP obtained by Chyb (1999), demonstrate that all members of the Drosophila TRPC family are regulated by fatty acids. In terms of phylogeny of signaling cascades, the questions arise whether regulation by fatty acids is a conserved feature and can also be found in other species or whether this mechanism is specific for flies. At present, it appears that most of the mammalian TRPC channels are activated by diacylglycerols (TRPC2, TRPC3, TRPC6, and TRPC7), whereas activation by fatty acids is a still unknown principle for mammalian TRPC channels. This is in line with the study in which PUFAs did not induce current signals in mock-transfected HEK293 cells, which endogenously express TRP channels. It therefore appears unlikely that endogenous TRP channels in HEK293 cells interfere with the activation of TRPγ by PUFAs (Jors, 2006).

The presence of endogenously expressed TRP channels in HEK293 cells is long known. For example, the first cloned TRP-homologous channel protein, TRPC1, has been cloned from HEK293 cells as well as TRPC3. In the meantime, RT-PCR experiments confirmed the expression of nearly all mammalian TRPC channels in HEK293 cells. The presence of natively expressed TRPC channels in HEK293 accounts for the receptor-induced responses recorded in mock-transfected cells and is discussed as contributing to the channel complexes formed in HEK293 cells heterologously expressing TRPC channels (Jors, 2006).

The work by Xu (1999) showed that heterologously expressed TRPγ forms a functional heteromeric channel complex together with TRPL. This ability would allow TRPγ and other TRP channel subunits to potentially generate a diversity of heteromeric channels, each with properties specifically tailored to a particular cellular function. This feature is a well known principle in other non-selective cation channel families, e.g. the cyclic nucleotide-gated channel subunits, the P2X channels, or the nicotinic acetylcholine channels. Results of fluorescence energy transfer assays and co-localization and co-immunoprecipitation studies have shown that co-expression of closely related proteins of the mammalian TRPC, TRPV, or TRPM families resulted in the formation of heteromeric channel complexes in HEK293 cells. Despite these results, co-expression of TRP channels in heterologous systems, such as HEK293 cells, had little impact on the functional properties of the channels studied. So far, only one study described that the co-expression of TRPC1 and TRPC5 resulted in a current-voltage relationship that differed from that of the homomeric channels. However, it has been shown that the heterologous expression of TRPV4 in HEK293 resulted in the down-regulation of a TRP-unrelated channel, the natively expressed volume-regulated anion channel. Any contribution of endogenous TRP channels of HEK293 cells to the regulation of the heterologously expressed TRPγ channels remains to be proven (Jors, 2006).

Whereas biochemical data argue in favor of a promiscuous interaction among TRP, TRPL, and TRPγ channels (Xu, 2000), the analysis of fly phototransduction demonstrated that TRPL channels alone translocated from the membrane of the rhabdomeres to intracellular membrane compartments in response to light. The fact that TRPL and TRPγ in the current study formed unitary homomeric channels activated by fatty acids may support the notion that these channels are involved in selective cellular signaling cascades, beyond their heteromeric function in phototransduction (Jors, 2006).

In summary, the current data show that TRPγ forms a Ca2+-permeable non-selective cation channel directly activated by polyunsaturated fatty acids generated from GPCR-dependent signaling pathways. Whereas a specific functional role of TRPγ in Drosophila phototransduction appears most attractive, its body-wide expression leads to the anticipation, however, of a more general role of TRPγ in the entire organism of the fruit fly (Jors, 2006).

Transient receptor potential-like channels are essential for calcium signaling and fluid transport in a Drosophila epithelium

Calcium signaling is an important mediator of neuropeptide-stimulated fluid transport by Drosophila Malpighian (renal) tubules. This study demonstrates the first epithelial role, in vivo, for members of the TRP family of calcium channels. RT-PCR revealed expression of trp, trpl, and trpγ in tubules. Use of antipeptide polyclonal antibodies for TRP, TRPL, and TRPγ showed expression of all three channels in type 1 (principal) cells in the tubule main segment. Neuropeptide (CAP2b)-stimulated fluid transport rates were significantly reduced in tubules from the trpl302 mutant and the trpl;trp double mutant, trpl302;trp343. However, a trp null, trp343, had no impact on stimulated fluid transport. Measurement of cytosolic calcium concentrations ([Ca2+]i) in tubule principal cells using an aequorin transgene in trp and trpl mutants showed a reduction in calcium responses in trpl302. Western blotting of tubule preparations from trp and trpl mutants revealed a correlation between TRPL levels and CAP2b-stimulated fluid transport and calcium signaling. Rescue of trpl302 with a trpl transgene under heat-shock control resulted in a stimulated fluid transport phenotype that was indistinguishable from wild-type tubules. Furthermore, restoration of normal stimulated rates of fluid transport by rescue of trpl302 was not compromised by introduction of the trp null, trp343. Thus, in an epithelial context, TRPL is sufficient for wild-type responses. Finally, a scaffolding component of the TRPL/TRP-signaling complex, INAD, is not expressed in tubules, suggesting that inaD is not essential for TRPL/TRP function in Drosophila tubules (MacPherson, 2005).

TRPγ channels are inhibited by cAMP and contribute to pacemaking in neurosecretory insect neurons

From a neuronal cDNA library of the cockroach Periplaneta americana a 3585-bp cDNA sequence encoding Periplaneta transient receptor potential γ (pTRPγ), a protein of 1194 amino acids showing 65% identity to the orthologous Drosophila channel protein dTRPγ was isolated. Heterologous expression of pTRPγ in HEK293 cells produced a constitutively active, non-selective cation channel with a Ca2+:Na+ permeability ratio of 2. In contrast to dTRPγ-mediated currents, pTRPγa currents were partially inhibited by 8-bromo-cAMP, and this effect was not mediated by protein kinase A (PKA) activation. pTRPγb, a truncated pTRPγ splice variant missing most of the C terminus, was insensitive to 8-bromo-cAMP. Thus, the critical cAMP-binding site seems to be located in the C-terminal part of pTRPγ, although there is no common cAMP-binding consensus sequence. While dTRPγ is only expressed in the photoreceptors, pTRPγ is expressed throughout the nervous system. In particular it is expressed in dorsal unpaired median (DUM) neurons. In these octopamine-releasing, neurosecretory cells a Ca2+ background current contributing to pacemaker activity was found to be up-regulated by the reduction of cAMP level. In addition, the Ca2+ background current was inhibited by LOE-908, 2-APB, and La3+, which similarly affected the pTRPγ current. It is thus proposed that the pTRPγ protein is involved in forming the channel passing the Ca2+ pacemaking background current in DUM neurons (Wicher, 2006).

TRPγ, 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, 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. This study identified such a subunit, TRPγ. TRPγ is highly enriched in photoreceptor cells and preferentially heteromultimerizes with TRPL in vitro and in vivo. The N-terminal domain of TRPγ dominantly suppressed the TRPL-dependent photoresponse, indicating that TRPgamma-TRPL heteromultimers contribute to the photoresponse. While TRPL and TRPγ homomultimers are constitutively active, this study demonstrated that TRPL-TRPγ heteromultimers form a regulated phospholipase C- (PLC-) stimulated channel (Xu, 2000).


Search PubMed for articles about Drosophila Trpγ

Akitake, B., Ren, Q., Boiko, N., Ni, J., Sokabe, T., Stockand, J. D., Eaton, B. A. and Montell, C. (2015). Coordination and fine motor control depend on Drosophila TRPγ. Nat Commun 6: 7288. PubMed ID: 26028119

Badsha, F., Kain, P., Prabhakar, S., Sundaram, S., Padinjat, R., Rodrigues, V. and Hasan, G. (2012). Mutants in Drosophila TRPC channels reduce olfactory sensitivity to carbon dioxide. PLoS One 7: e49848. PubMed ID: 23185459

Chiang, A. S., Blum, A., Barditch, J., Chen, Y. H., Chiu, S. L., Regulski, M., Armstrong, J. D., Tully, T. and Dubnau, J. (2004). radish encodes a phospholipase-A2 and defines a neural circuit involved in anesthesia-resistant memory. Curr Biol 14: 263-272. PubMed ID: 14972677

Chyb, S., Raghu, P. and Hardie, R. C. (1999). Polyunsaturated fatty acids activate the Drosophila light-sensitive channels TRP and TRPL. Nature 397: 255-259. PubMed ID: 9930700

Jors, S., Kazanski, V., Foik, A., Krautwurst, D. and Harteneck, C. (2006). Receptor-induced activation of Drosophila TRP γ by polyunsaturated fatty acids. J Biol Chem 281: 29693-29702. PubMed ID: 16901908

Kain, P., Chakraborty, T. S., Sundaram, S., Siddiqi, O., Rodrigues, V. and Hasan, G. (2008). Reduced odor responses from antennal neurons of G(q)alpha, phospholipase Cbeta, and rdgA mutants in Drosophila support a role for a phospholipid intermediate in insect olfactory transduction. J Neurosci 28: 4745-4755. PubMed ID: 18448651

MacPherson, M. R., Pollock, V. P., Kean, L., Southall, T. D., Giannakou, M. E., Broderick, K. E., Dow, J. A., Hardie, R. C. and Davies, S. A. (2005). Transient receptor potential-like channels are essential for calcium signaling and fluid transport in a Drosophila epithelium. Genetics 169: 1541-1552. PubMed ID: 15695363

Shortridge, R. D., Yoon, J., Lending, C. R., Bloomquist, B. T., Perdew, M. H. and Pak, W. L. (1991). A Drosophila phospholipase C gene that is expressed in the central nervous system. J Biol Chem 266: 12474-12480. PubMed ID: 2061323

Wicher, D., Agricola, H. J., Schonherr, R., Heinemann, S. H. and Derst, C. (2006). TRPγ channels are inhibited by cAMP and contribute to pacemaking in neurosecretory insect neurons. J Biol Chem 281: 3227-3236. PubMed ID: 16319060

Xu, X. Z., Chien, F., Butler, A., Salkoff, L. and Montell, C. (2000). TRPγ, a Drosophila TRP-related subunit, forms a regulated cation channel with TRPL. Neuron 26: 647-657. PubMed ID: 10896160

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date revised: 14 December 2022

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