ripped pocket: Biological Overview | References
Gene name - ripped pocket
Cytological map position - 82C5-82C5
Function - channel
Keywords - DEG/ENaC channel family member - reduces depolarization and Hh signal transduction in the wing disc - ovaries - blocking Ripped pocket leads to defects in force generation during dorsal closure via failure of actomyosin structures - functions in the PNS to regulate behavioral responses to touch and in the formation of the actin-rich sensory filopodia
Symbol - rpk
FlyBase ID: FBgn0022981
Genetic map position - chr3R:4,645,981-4,648,884
Classification - Amiloride-sensitive sodium channel
Cellular location - surface transmembrane
While the membrane potential of cells has been shown to be patterned in some tissues, specific roles for membrane potential in regulating signalling pathways that function during development are still being established. In the Drosophila wing imaginal disc, Hedgehog (Hh) from posterior cells activates a signalling pathway in anterior cells near the boundary which is necessary for boundary maintenance. This study shows that membrane potential is patterned in the wing disc. Anterior cells near the boundary, where Hh signalling is most active, are more depolarized than posterior cells across the boundary. Elevated expression of the ENaC channel Ripped Pocket (Rpk), observed in these anterior cells, requires Hh. Antagonizing Rpk reduces depolarization and Hh signal transduction. Using genetic and optogenetic manipulations, in both the wing disc and the salivary gland, it was shown that membrane depolarization promotes membrane localization of Smoothened and augments Hh signalling, independently of Patched. Thus, membrane depolarization and Hh-dependent signalling mutually reinforce each other in cells immediately anterior to the compartment boundary (Emmons-Bell, 2021).
Initially discovered for its role in regulating segment polarity in Drosophila, Hh signalling has since been implicated in a multitude of developmental processes. Among the best characterized is the signalling between two populations of cells that make up the Drosophila wing imaginal disc, the larval primordium of the adult wing and thorax. The wing disc consists of two compartments of lineage-restricted cells separated by a smooth boundary. Posterior (P) cells make the morphogen Hedgehog, which binds to its receptor Patched (Ptc), which is expressed exclusively in anterior (A) cells. Hh has a relatively short range either because of its limited diffusion, or because it is taken up by nearby target cells via filopodia-like protrusions known as cytonemes. Hh alleviates the repressive effect of Ptc on the seven-transmembrane protein Smoothened (Smo) in A cells near the boundary, initiating a signalling cascade that culminates in the stabilization of the activator form of the transcription factor Cubitus interruptus (Ci), and expression of target genes such as the long-range morphogen Dpp. In turn, Dpp regulates imaginal disc patterning and growth in both compartments (Emmons-Bell, 2021).
While the role of cell-cell interactions, diffusible morphogens and even mechanical forces have been studied in regulating the growth and patterning of the wing disc, relatively little attention has been paid to another cellular parameter, membrane potential or Vmem. Vmem is determined by the relative concentrations of different species of ions across the cell membrane, as well as the permeability of the membrane to each of these ions. These parameters are influenced by the abundance and permeability of ion channels, the activity of pumps, and gap junctions. While changes in Vmem have been studied most extensively in excitable cells, there is increasing evidence that the Vmem of all cells, including epithelial cells, can vary depending on cell-cycle status and differentiation status. Mutations in genes encoding ion channels in humans ('channelopathies') can result in congenital malformations. Similarly, experimental manipulation of ion channel permeability can cause developmental abnormalities in mice as well as in Drosophila. Only more recently has evidence emerged that Vmem can be patterned during normal development. Using fluorescent reporters of membrane potential, it has been shown that specific cells during Xenopus gastrulation and Drosophila oogenesis appear more depolarized than neighbouring cells. A recent study established that cells in the vertebrate limb mesenchyme become more depolarized as they differentiate into chondrocytes, and that this depolarization is essential for the expression of genes necessary for chondrocyte fate. However, in many of these cases, the relationship between changes in Vmem and specific pathways that regulate developmental patterning have not been established (Emmons-Bell, 2021).
This study investigated the patterning of Vmem during wing disc development and showed that the regulation of Vmem has an important role in regulating Hh signalling. The cells immediately anterior to the compartment boundary, a zone of active Hh signalling, are more depolarized than surrounding cells, and Hh signalling and depolarized Vmem mutually reinforce each other. This results in an abrupt change in Vmem at the compartment boundary (Emmons-Bell, 2021).
This study shows that Vmem is patterned in a spatiotemporal manner during development of the wing disc of Drosophila and that it regulates Hedgehog signalling at the compartment boundary. First, it was shown that cells immediately anterior to the compartment boundary are relatively more depolarized than cells elsewhere in the wing pouch. This region coincides with the A cells where Hh signalling is most active, as evidenced by upregulation of Ptc. Second, the expression of at least two regulators of Vmem, the ENaC channel Rpk and the alpha subunit of the Na+/K+ ATPase were shown to be expressed at higher levels in this same portion of the disc. Third, by altering Hh signalling, this study demonstrated that the expression of both Rpk and ATPα is increased in cells with increased Hh signalling. Fourth, by manipulating Hh signalling in the disc and using optogenetic methods, both in the salivary gland and wing disc, it was shown that membrane depolarization promotes Hh signalling as assessed by increased membrane localization of Smo, and expression of the target gene ptc. Thus, Hh-induced signalling and membrane depolarization appear to mutually reinforce each other and thus contribute to the mechanisms that maintain the segregation of A and P cells at the compartment boundary (Emmons-Bell, 2021).
Two regions of increased DiBAC fluorescence were observed in the wing imaginal disc. No obvious upregulation of Rpk and ATPα was observed in other discs, and therefore, these studies have focused on the region immediately anterior to the A-P compartment boundary in the wing disc. In the late L3 wing disc, a region of increased DiBAC fluorescence was observed in the A compartment in the vicinity of the D-V boundary. This corresponds to a 'zone of non-proliferating cells' (ZNC). Interestingly, the ZNC is different in the two compartments. In the A compartment, two rows of cells are arrested in G2 while in the P compartment, a single row of cells is arrested in G1. The observation of increased DiBAC fluorescence in the DV boundary of only the A compartment is consistent with previous reports that cells become increasingly depolarized as they traverse S-phase and enter G2. In contrast, cells in G1 are thought to be more hyperpolarized. Additionally, increased expression of the ENaC channel Rpk was observed in two rows of cells at the D-V boundary in the anterior compartment, indicating that increased expression of Rpk could contribute to the depolarization observed in those cells. It is noted, however, that the increased DiBAC fluorescence in these cells was not entirely eliminated by exposing discs to amiloride, indicating that other factors are also likely to contribute (Emmons-Bell, 2021).
These data are consistent with a model where membrane depolarization and Hh-induced signalling mutually reinforce each other in the cells immediately anterior to the compartment boundary. Both membrane depolarization and the presence of Hh seem necessary for normal levels of activation of the Hh signalling pathway in this region; neither alone is sufficient. First, it was shown that Hh signalling promotes membrane depolarization. It was also shown that the expression of Rpk just anterior to the A-P compartment boundary is dependent upon Hh signalling. Elevated Rpk expression is not observed when a hhts allele is shifted to the restrictive temperature, and cells become more depolarized when Hh signalling is constitutively activated through expression of the ci3m allele. Previously published microarray data suggest that Rpk as well as another ENaC family channel Ppk29 are both enriched in cells that also express ptc. However, there is no antibody to assess Ppk29 expression currently. The sensitivity of the depolarization to amiloride indicates that these and other ENaC channels make an important contribution to the membrane depolarization (Emmons-Bell, 2021).
Second, this study has shown that the depolarization increases Hh signalling. The early stages of Hh signalling are still incompletely understood. Hh is thought to bind to a complex of proteins that includes Ptc together with either Ihog or Boi. This alleviates an inhibitory effect on Smo, possibly by enabling its access to specific membrane sterols. Interestingly, it has recently been proposed that Ptc might function in its inhibitory capacity by a chemiosmotic mechanism where it functions as a Na+ channel. An early outcome of Smo activation is its localization to the membrane where its C-terminal tail becomes phosphorylated and its ubiquitylation and internalization are prevented. By manipulating channel expression in the wing disc, and by optogenetic experiments in both the salivary gland and wing disc, this study has shown that membrane depolarization can promote Hh signalling as assessed by increased Smo membrane localization and increased expression of the target gene ptc. The time course of Smo activation is relatively rapid (over minutes) and is therefore unlikely to require new transcription and translation. In the P compartment, membrane Smo levels are elevated likely because of the complete absence of Ptc, and some downstream components of the Hh signalling pathway are known to be activated. However, since Ci is not expressed in P cells, target gene expression is not induced. In the cells just anterior to the boundary, the partial inhibition of Ptc by Hh together with membrane depolarization seem to combine to achieve similar levels of Smo membrane localization. More anteriorly, the absence of this mutually reinforcing mechanism appears to result in Smo internalization (Emmons-Bell, 2021).
The experiments do not point to a single mechanism by which depolarization promotes Hh signalling. It is possible that depolarization results in increased Ca2+ levels by opening Ca2+channels at the plasma membrane or by promoting release from intracellular sources (e.g. the ER or mitochondria). Indeed, there is evidence that Ca2+ entry into the primary cilium promotes Hh signalling, and recent work shows that targets of Sonic Hedgehog (Shh) signalling during mammalian development is augmented by Ca2+ influx. A second possibility is that membrane depolarization could, by a variety of mechanisms, activate the kinases that phosphorylate the C-terminal tail of Smo and maintain it at the plasma membrane in an activated state. Depolarization could also impact electrostatic interactions at the membrane that make the localization of Smo at the membrane more favourable. Since Rpk and ATPα are expressed at higher levels in the cells that receive Hh, which have been postulated to make synapse-like projections with cells that produce Hh, it is conceivable that these channels could modulate synapse function. Additionally, while this work was under review, it has been reported that reducing glycolysis depletes ATP levels and results in depolarization in the wing imaginal disc, reducing the uptake of Hh pathway inhibitors and stabilizing Smo at the cell membrane (Spannl, 2020). Importantly, all these mechanisms are not mutually exclusive and their roles in Hh signalling are avenues for future research (Emmons-Bell, 2021).
It is now generally accepted that both cell-cell signalling and mechanical forces have important roles in cell fate specification and morphogenesis. This work adds to a growing body of literature suggesting that changes in Vmem, a relatively understudied parameter, may also have important roles in development. Integrating such biophysical inputs with information about gene expression and gene regulation will lead to a more holistic understanding of development and morphogenesis (Emmons-Bell, 2021).
This study has identified the first known function for the class II and the class III md neurons. These neurons showed physiological responses to force, their output was required for touch responses, and their optogenetic activation was sufficient to generate behavioral responses that resembled the behavioral responses to touch. This study focused on the class III neurons and their actin-rich sensory filopodia. The results indicated that the filopodia contribute to both physiological (Ca2+ responses) and behavioral responses to gentle touch (Schotthofer, 2020).
Time-lapse imaging indicated that sensory filopodia in class III neurons are very dynamic during the early third-instar stage, which has interesting similarity to the actin-mediated spine motility known to occur at immature synapses. Interestingly, sensory filopodia are quite stable in late wandering third instar in comparison to the early third instar, thus the dynamic nature of these filopodia may also be related to a developmental process in which the overall mechanical sensitivity of the neuron is tuned as the dendritic arbor scales during the profound growth period of the third instar (Schotthofer, 2020).
An interesting possibility is that the actin-rich structures may hold mechanosensitive channels in a similar fashion to the stereovilli of hair cells in the vertebrate inner ear. If this is the case, growth and relaxation of the sensory filopoida may be important for fine-tuning the plasma membrane tension that is needed for proper mechanotransduction (Schotthofer, 2020).
To begin to elucidate the molecular mechanisms by which class III neurons sense external signals, a comprehensive RNAi screen was performed for channel genes that were required for gentle touch responses. Ion channel gene families previously implicated in mechanotransduction (TRP, DEG/ENaC, and NMDARs) were identified in this screen. Although it is tempting to speculate from these results that one or more of the channel subunits isolated from the screen might be mechanically sensitive, further experimentation will be needed to adequately test this hypothesis (Schotthofer, 2020).
This study focused on the rpk gene, which is a member of the DEG/ENaC family. Formal genetic proof is provided that this gene is essential for gentle touch responses in the Drosophila larvae. Interestingly, the DEG/ENaC subunit encoded by the pickpocket locus is required for mechanical nociception. This function is consistent with the very specific expression pattern of the ppk gene in class IV neurons. Although ppk is not required for gentle touch responses, ppk-GAL4 is weakly expressed in the class III neurons. Thus, although not essential for gentle touch responses, PPK represents a potential heteromeric partner subunit for RPK in these cells (Schotthofer, 2020).
The results suggest that the identified ion channel subunits contribute to the stability of the sensory filopodia. Prior results implicated calcium/calmodulin-dependent protein kinase II (CaMKII) in sensory filopodia growth; however, the source of calcium ions that activate CaMKII in the class III neurons is unknown. The NOMPC channel and the NMDARs identified in this study represent good candidates for providing mechanically activated Ca2+ entry that could drive the activation of CaMKII. Local activation of Rho family GTPases in postsynaptic dendritic spines can also be driven by CaMKII. So the reduced filopodia formation seen in ion channel knockdown experiments may also involve pathways utilizing the same Rho family members that were manipulated in this study. When dendritic morphology was manipulated by expressing wild-type and dominant-negative forms of Rho family GTPases in class III neurons, both filopodia number and behavioral sensitivity to gentle touch were dramatically changed (Schotthofer, 2020).
In conclusion, actin-rich dendritic sensory filopodia on nonciliated Drosophila class III neurons represent sensory organelles that play an important role in efficient somatosensory mechanosensation. An ensemble of ion channels interacts with these sensory filopodia in generating touch responses. Importantly, F-actin has been implicated in mechanosensory responses of mammalian somatosensory neurons and actin-rich protrusions have been observed on mechanosensory Merkel cells. An interesting possibility is that mammalian somatosensation relies upon structures that are evolutionarily homologous to the actin-rich sensory filopodia of the Drosophila class III neurons. Thus, the further study of these structures may shed important light on the mechanisms of mammalian touch sensation (Schotthofer, 2020).
Ion channels contribute to the regulation of dorsal closure in Drosophila, a model system for cell sheet morphogenesis. Ca2+ was found to be sufficient to cause cell contraction in dorsal closure tissues, as UV-mediated release of caged Ca2+ leads to cell contraction. Furthermore, endogenous Ca2+ fluxes correlate with cell contraction in the amnioserosa (AS) during closure, whereas the chelation of Ca2+ slows closure. Microinjection of high concentrations of the peptide GsMTx4, which is a specific modulator of mechanically gated ion channel function, causes increases in cytoplasmic free Ca2+ and actomyosin contractility and, in the long term, blocks closure in a dose-dependent manner. Two channel subunits, ripped pocket and dtrpA1 (TrpA1), were identified that play a role in closure and other morphogenetic events. Blocking channels leads to defects in force generation via failure of actomyosin structures, and impairs the ability of tissues to regulate forces in response to laser microsurgery. These results point to a key role for ion channels in closure, and suggest a mechanism for the coordination of force-producing cell behaviors across the embryo (Hunter, 2014).
These data provide evidence that ion channels function in closure to regulate ion flux in individual cells in the AS and leading edge (LE), leading to Ca2+-dependent cell contractility. Intracellular ion flux via mechanically gated ion channels (MGCs) can promote cytoskeletal and junction organization in cell culture. The findings that localization of the Ca2+ reporter C2:GFP correlates with AS cell contraction and that elevated Ca2+ induces AS cell contraction (via uncaging NP EGTA AM or spontaneous flashes) support a role for Ca2+-dependent contractility in closure. Nevertheless, the observed correlation between perimeter shortening (contraction) and increases in free Ca2+ is by no means perfect. It is hypothesized that there are several reasons for this lack of tight correlation. First, although the data suggest that Ca2+ plays a role in regulating actomyosin contractility, there are other regulators of this important process, and small GTPases (especially Rac and Rho) are sure to play a regulatory role. Ca2+ signaling must be integrated into the context of other signaling pathways and programs of gene expression regulating morphogenesis. Second, individual cell behavior must be considered in the context of the AS cell sheet, in which the behavior of a cell perimeter is profoundly influenced by the behavior of the cells to which it is attached. It is possible that passive perimeter shortening on one side of a given cell is actively driven by contractility in its neighbor. Finally, two-dimensional changes in cell shape, as observed in a given optical section or series of optical sections, must be considered in the context of the three-dimensional nature of cells. It is hypothesized that cell volume does not fluctuate rapidly because of the relative incompressibility of cellular constituents and because the cell does not rapidly lose or gain volume. Thus, cell volume acts as a buffer and changes in crosssectional area (e.g., measured at the level of junctional belts) may be the consequence of contractile activities functioning elsewhere in the cell. Complete understanding of how MGCs and Ca2+-mediated contraction are integrated into cellular homeostasis and morphogenesis requires a more complete picture of how other signaling pathways contribute to changes in cell shape. Moreover, it will require more complete imaging sets, with higher temporal and spatial resolution, of the three-dimensional changes that occur during morphogenesis, even in relatively simple morphogenetic movements such as dorsal closure. The advent of new biosensors and high-speed imaging techniques place the technologies required for such investigations of morphogenesis within the realm of possibility (Hunter, 2014).
GsMTx4 is the most specific pharmacological reagent for manipulating MGC activity in vitro and in vivo, and this study reports its use during Drosophila embryogenesis. Acute, bimodal effects of GsMTx4 on closure are consistent with the presence of MGCs that ultimately pass Ca2+ ions. At the microM concentrations of GsMTx4 experienced by cells at or near the site of injection, increases in cellular free Ca2+ are followed by constriction over the course of tens of seconds. By contrast, the long-term effects of low concentrations of GsMTx4, which cells experience after the bolus of peptide diffuses away from the injection site, appear to be inhibition of closure via the failure of key actomyosin structures and activities. It is hypothesized that GsMTx4 affects MGC activity by modifying the thickness or curvature of the lipid bilayer in which these channels are embedded, consistent with known mechanisms of GsMTx4 action. Studies in cell culture demonstrate that loss of MGC function by pharmacological inhibitors or targeted mutations in channel subunits leads to defects in actomyosin contractile behaviors. Nevertheless, it cannot be ruled out that possible indirect effects of MGC inhibition obscure specific and direct long-term effects of MGC inhibition (e.g. the effect of membrane thickness and curvature on non-MGC membrane proteins during development or secondary consequences of inhibiting RPK and dTRPA1) (Hunter, 2014).
Long-term phenotypes due to GsMTx4-mediated MGC inhibition are recapitulated by RNAi expression or mutational analysis that disrupt the function of specific channels. Congenital loss of channel expression is a long-term effect, and disrupting expression of rpk or dtrpA1 in embryos leads to closure defects. Discrepancies in phenotypes may be the consequence of multiple channels functioning in closure or to differences in the timing and pattern of knockdown or inhibition. Whereas in the current experiments, the expression knockdown of a single channel subunit tissue specifically (via RNAi) or in the embryo as a whole prior to closure (via mutant allele), an advantage of pharmacological inhibitors is acute delivery before or during closure. Indeed, phenotypes were observed consistent with genetic knockdown when GsMTx4 was knocked down prior to closure: defects in AS shape, canthus and purse string formation and failure to close. It is speculated that the embryo can compensate for the congenital loss of a single channel subunit (as in the case of dtrpA1) in ways not possible when drug is applied acutely or when RNAi knocks down channel function (less acutely than the drug, more acutely than inherited mutant alleles). The regulation of contractility via ion channels during closure appears to be both cell-autonomous and non-cell-autonomous (Hunter, 2014).
Specifically, the loss of leading edge cell elongation and their purse strings when channel subunits are targeted in the AS indicates that robust channel activity in the AS is required for normal cell shape changes in both the AS (i.e. cell-autonomous) and in leading edge cells (i.e. non-cell-autonomous). Actomyosin contractility during closure can act non-cell-autonomously, implicating positive reinforcement of force-producing activities or structures between and within embryonic tissues: clones of cells expressing myosin because of a transgenic mosaic effect contract (cell-autonomous effect) but stretch neighboring cells (non-cell-autonomous effect). It is hypothesized that channel activity contributes to tension at a single-cell level in the AS, and that tension in the AS, exerted on the LE, is required for wild-type actomyosin-dependent structures and cell shapes in the leading edge of the LE (Hunter, 2014).
Verification of a mechanical circuit(s) regulated by MGCs requires that which channels are involved can be unequivocally established, and the gating mechanisms of each channel(s) be determined in the embryonic epithelia. The sensitivity of DEG/ENaC and TRPA1 homologs to applied force has been studied in other systems, but is unknown for dorsal closure tissues. Future studies should include electrophysiological recordings, but such methods have not yet been developed for analysis of Drosophila embryonic epithelial cells. These studies could be key for understanding how a Na+-permeable channel (RPK) contributes to Ca2+ flux. Although its ability to conduct Ca2+ or associate with Ca2+ channel subunits is unknown, RPK is involved in Ca2+-dependent processes such as Drosophila oocyte activation and the response to gentle touch in larvae. This study implicates ion flux and MGCs in the molecular mechanisms that regulate closure. Force sensing by MGCs could constitute a rapid means of affecting cell behaviors in order to adapt to acute changes during closure. For example, at the level of apical junctions, individual AS cells change shape dramatically, whereas the overall area of the AS decreases slowly and monotonically. Based on the current observations, it is hypothesized that MGCs function in a mechanical circuit(s) to coordinate forces across the embryo. Similar feedback loops are proposed for the oscillatory behavior of other mechanically coupled, contractile cells. Given that morphogenesis throughout Drosophila development requires the assembly and regulation of force-producing structures, it will be interesting to determine how other morphogenetic processes are affected by channel inhibition (Hunter, 2014).
The molecular and functional characteristics of Ripped pocked (Rpk) were examined. Expression of an epitope-tagged Rpk construct in COS-7 cells generated a 73-kD glycoprotein that can be deglycosylated to its predicted molecular mass of 65 kD. On Northern analysis, an rpk probe detected transcripts in only embryonic and adult RNA, where a major 3.4-kb transcript was observed, as well as two smaller less-abundant messages (Adams, 1998).
To determine the embryonic expression pattern of rpk transcripts, in situ hybridization to whole mount embryos was performed using an antisense rpk probe. In contrast to ppk transcripts, rpk transcripts are detected in early stage (0-3 h) embryos, but are not present in later stages of embryogenesis. Furthermore, in early stage embryos, rpk transcripts are not localized to a specific embryonic region or cell type. In Drosophila embryos, zygotic transcription does not initiate until the third hour of development. Because rpk mRNA is detected in embryos before the initiation of zygotic transcription, this result suggests that embryonic rpk message is of maternal origin, and that Rpk may play a role in early development (Adams, 1998).
When expressed in Xenopus oocytes, Rpk generates small whole cell Na+ currents that are reversibly blocked by amiloride. Rpk is impermeable to K+, as shown by the elimination of inward current when external Na+ is replaced with K+. Thus, in contrast to Ppk, Rpk forms functional ion channels by itself (Adams, 1998).
In several C. elegans degenerins, mutation of a specific residue near M2, the 'Deg' mutation, causes a dominant form of neurodegeneration suggestive of constitutive ion channel activity. Similarly, BNC1 containing a Deg mutation (BNC1G430V) is activated, producing much larger currents in Xenopus oocytes. To learn whether Rpk could also be activated by the Deg mutation, a valine residue was incorporated at the appropriate position (residue 524). Like wild-type Rpk, RpkA524V generates Na+-selective currents that are reversibly inhibited by amiloride. However, RpkA524V currents are 20-50 times larger than wild-type Rpk currents. This indicates that the Deg mutation activates Rpk. RpkA524V is slightly more permeable to Li+ than Na+ but was impermeable to K+. RpkA524V is significantly more sensitive to amiloride than wild-type Rpk. Gadolinium, an inhibitor of mechanosensation and some stretch-activated channels, also reversibly inhibits RpkA524V current (Adams, 1998).
Individual DEG/ENaC proteins are subunits that form homo- or hetero-multimeric ion channels. Because DEG/ ENaC proteins with Deg mutations produce a genetically dominant phenotype in C. elegans, it is thought that the Deg mutation in one or a few subunits might activate the channel complex, producing larger currents. The hypothesis that the Deg mutation is dominant at the molecular level was tested by asking if channels composed of both wild-type Rpk and RpkA524V would generate small or large Na+ currents. Coexpression of Rpk and RpkA524V generate large Na+ currents that are similar in size to those generated by RpkA524V alone. However, the amiloride sensitivity of the current, was similar to that generated by wild-type Rpk alone. These observations indicate that the increase in current amplitude depends on RpkA524V, and the low amiloride sensitivity depends on wild-type Rpk. Thus, the data suggest that at least two subunits combine to produce multimeric channels, that the A524V mutation dominantly activates the channel, and that Ala524 dominantly determines amiloride sensitivity. Gadolinium also inhibits wild-type Rpk current, and gadolinium sensitivity is not significantly altered by the presence of wild-type Rpk in a complex with RpkA524V. Coexpression of Ppk with Rpk or RpkA524V does not significantly alter the amount, ionic selectivity, or amiloride sensitivity of Rpk or RpkA524V current. Thus, it appears that Ppk and Rpk are not subunits of the same ion channel but likely have distinct physiological roles (Adams, 1998).
Search PubMed for articles about Drosophila Ripped pocket
Adams, C. M., Anderson, M. G., Motto, D. G., Price, M. P., Johnson, W. A. and Welsh, M. J. (1998). Ripped pocket and Pickpocket, novel Drosophila DEG/ENaC subunits expressed in early development and in mechanosensory neurons. J. Cell Biol. 140: 143-152. 9425162
Emmons-Bell, M. and Hariharan, I. K. (2021). Membrane potential regulates Hedgehog signalling in the Drosophila wing imaginal disc. EMBO Rep: e51861. PubMed ID: 33629503
Hunter, G. L., Crawford, J. M., Genkins, J. Z. and Kiehart, D. P. (2014). Ion channels contribute to the regulation of cell sheet forces during Drosophila dorsal closure. Development 141(2): 325-34. PubMed ID: 24306105
Schotthofer, S. K. and Bohrmann, J. (2020). Analysing bioelectrical phenomena in the Drosophila ovary with genetic tools: tissue-specific expression of sensors for membrane potential and intracellular pH, and RNAi-knockdown of mechanisms involved in ion exchange. BMC Dev Biol 20(1): 15. PubMed ID: 32635900
Spannl, S., Buhl, T., Nellas, I., Zeidan, S. A., Iyer, K. V., Khaliullina, H., Schultz, C., Nadler, A., Dye, N. A. and Eaton, S. (2020). Glycolysis regulates Hedgehog signalling via the plasma membrane potential. EMBO J 39(21): e101767. PubMed ID: 33021744
Tsubouchi, A., Caldwell, J. C. and Tracey, W. D. (2012). Dendritic filopodia, Ripped Pocket, NOMPC, and NMDARs contribute to the sense of touch in Drosophila larvae. Curr Biol 22(22): 2124-2134. PubMed ID: 23103192
date revised: 13 May 2022
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