Balboa Binds to Pickpocket In Vivo and Is Required for Mechanical Nociception in Drosophila Larvae

Abstract

Summary The Drosophila gene pickpocket (ppk) encodes an ion channel subunit of the Degenerin/Epithelial (DEG/ENaC) family [1]. PPK is specifically expressed in nociceptive, Class IV multidendritic (md) neurons and is functionally required for mechanical nociception responses [2, 3]. In this study, in a genome-wide genetic screen for other ion channel subunits required for mechanical nociception, we identify a gene that we name balboa (a.k.a. CG8546, ppk26) [4]. Interestingly, the balboa locus encodes a DEG/ENaC ion channel subunit highly similar in amino acid sequence to PPK [5]. Moreover, laser-capture isolation of RNA from larval neurons and microarray analyses reveal that balboa is also highly enriched in nociceptive neurons. The requirement for Balboa and PPK in mechanical nociception behaviors and their specific expression in larval nociceptors led us to hypothesize that these DEG/ENaC subunits form an ion channel complex in vivo. In nociceptive neurons Balboa∷GFP proteins distribute uniformly throughout dendrites but remarkably localize to discrete foci when ectopically expressed in other neuron subtypes (where PPK is not expressed). Indeed, ectopically co-expressing ppk transforms this punctate Balboa∷GFP expression pattern to the uniform distribution observed in its native cell-type. Furthermore, ppk-RNAi in Class IV neurons alters the broad Balboa∷GFP pattern to a punctate distribution. Interestingly, this interaction is mutually codependent as balboa-RNAi eliminates Venus∷PPK from the sensory dendrites of nociceptors. Finally, using a GFP-reconstitution approach in transgenic larvae, we directly detect in vivo physical interactions among PPK and Balboa subunits. Combined, our results indicate a critical mechanical nociception function for heteromeric PPK and Balboa channels in vivo.

Tissue mechanics govern the rapidly adapting and symmetrical response to touch

“…A transient receptor potential protein performs the role in a fifth class of mechanoreceptor that functions as a texture sensor (9,(17)(18)(19)(20). Both kinds of ion channels also contribute to touch and pain sensation in Drosophila larvae (9,11,14,(21)(22)(23)(24)(25). Collectively, these observations suggest that no single class of proteins is responsible for forming sensory MeT channels and that the properties of rapid adaptation and symmetrical on and off responses are not uniquely linked to a single class of channel proteins.…”

In vivo dendrite regeneration after injury is different from dendrite development

“…4A-D). Moreover, in class IV neurons, we found that the DEG/ ENaC channels ppk1 and ppk26, which are necessary for the function of the dendrites in mechanical nociception (Zhong et al 2010;Gorczyca et al 2014;Guo et al 2014;Mauthner et al 2014), are properly trafficked into regenerated dendrites and properly excluded from the axon (Fig. 6B,C).…”

“…The DEG/ENaC channels ppk1 and ppk26, which are necessary for mechanical nociception, are normally specifically expressed by class IV da neurons (Zhong et al 2010;Gorczyca et al 2014;Guo et al 2014;Mauthner et al 2014). We showed above that these proteins are properly trafficked into regenerated dendrites and excluded from axons, but are they still expressed by the correct neuronal subtype?…”

Role of kinesin-1–based microtubule sliding in Drosophila nervous system development

“…Class IV da neurons have been reported to detect noxious thermal (47) and mechanical stimuli (48,49), triggering larvae to engage in escape behaviors to evade predators such as parasitoid wasps (50,51). Previous studies have shown that noxious mechanical stimulation to the middle segments of third-instar larvae results in a reproducible, dorsalventral rolling escape behavior (51).…”