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  1. Mahdieh, Nejat (Ed.)

    Johnston’s organ, theDrosophilaauditory organ, is anatomically very different from the mammalian organ of Corti. However, recent evidence indicates significant cellular and molecular similarities exist between vertebrate and invertebrate hearing, suggesting thatDrosophilamay be a useful platform to determine the function of the many mammalian deafness genes whose underlying biological mechanisms are poorly characterized. Our goal was a comprehensive screen of all known orthologues of mammalian deafness genes in the fruit fly to better understand conservation of hearing mechanisms between the insect and the fly and ultimately gain insight into human hereditary deafness. We used bioinformatic comparisons to screen previously reported human and mouse deafness genes and found that 156 of them have orthologues inDrosophila melanogaster. We used fluorescent imaging of T2A-GAL4 gene trap and GFP or YFP fluorescent protein trap lines for 54 of theDrosophilagenes and found 38 to be expressed in different cell types in Johnston’s organ. We phenotypically characterized the function of strong loss-of-function mutants in three genes expressed in Johnston’s organ (Cad99C,Msp-300, andKoi) using a courtship assay and electrophysiological recordings of sound-evoked potentials.Cad99CandKoiwere found to have significant courtship defects. However, when we tested these genes for electrophysiological defects in hearing response, we did not see a significant difference suggesting the courtship defects were not caused by hearing deficiencies. Furthermore, we used a UAS/RNAi approach to test the function of seven genes and found two additional genes,CG5921andMyo10a, that gave a statistically significant delay in courtship but not in sound-evoked potentials. Our results suggest that many mammalian deafness genes haveDrosophilahomologues expressed in the Johnston’s organ, but that their requirement for hearing may not necessarily be the same as in mammals.

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    Free, publicly-accessible full text available February 27, 2025
  2. We show here thatmir-279/996are absolutely essential for development and function of Johnston's organ (JO), the primary proprioceptive and auditory organ inDrosophila. Their deletion results in highly aberrant cell fate determination, including loss of scolopale cells and ectopic neurons, and mutants are electrophysiologically deaf. In vivo activity sensors and mosaic analyses indicate that these seed-related miRNAs function autonomously to suppress neural fate in nonneuronal cells. Finally, genetic interactions pinpoint two neural targets (elavandinsensible) that underlie miRNA mutant JO phenotypes. This work uncovers how critical post-transcriptional regulation of specific miRNA targets governs cell specification and function of the auditory system.

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    Free, publicly-accessible full text available November 1, 2024
  3. Introduction

    Cilia biogenesis relies on intraflagellar transport (IFT), a conserved transport mechanism which functions bi-directionally to bring protein complexes to the growing ciliary tip and recycle signaling and transport proteins between the cilium and cell body. InDrosophila, anterograde IFT is critical for assembly of sensory cilia in the neurons of both chordotonal (ch) organs, which have relatively long ciliary axonemes, and external sensory (es) organs, which have short axonemal segments with microtubules in distal sensory segments forming non-axonemal bundles. We previously isolated thebeethoven(btv) mutant in a mutagenesis screen for auditory mutants. Although manybtvmutant flies are deaf, some retain a small residual auditory function as determined both by behavior and by auditory electrophysiology.


    Here we molecularly characterize thebtvgene and demonstrate that it encodes the IFT-associated dynein-2 heavy chain Dync2h1. We also describe morphological changes in Johnston’s organ as flies age to 30 days, and we find that morphological and electrophysiological phenotypes in this ch organ ofbtvmutants become more severe with age. We show that NompB protein, encoding the conserved IFT88 protein, an IFT complex B component, fails to be cleared from chordotonal cilia inbtvmutants, instead accumulating in the distorted cilia. In macrochaete bristles, a class of es organ,btvmutants show a 50% reduction in mechanoreceptor potentials.


    Thus, thebtv-encoded Dync2h1 functions as the retrograde IFT motor in the assembly of long ciliary axonemes in ch organs and is also important for normal function of the short ciliary axonemes in es organs.

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    Free, publicly-accessible full text available September 22, 2024
  4. Abstract

    The fruit flyDrosophila melanogasterhas provided important insights into how sensory information is transduced by transient receptor potential (TRP) channels in the peripheral nervous system (PNS). However, TRP channels alone have not been able to completely model mechanosensitive transduction in mechanoreceptive chordotonal neurons (CNs). Here, we show that, in addition to TRP channels, the sole voltage-gated sodium channel (NaV) inDrosophila, Para, is localized to the dendrites of CNs. Para is localized to the distal tip of the dendrites in all CNs, from embryos to adults, and is colocalized with the mechanosensitive TRP channels No mechanoreceptor potential C (NompC) and Inactive/Nanchung (Iav/Nan). Para localization also demarcates spike initiation zones (SIZs) in axons and the dendritic localization of Para is indicative of a likely dendritic SIZ in fly CNs. Para is not present in the dendrites of other peripheral sensory neurons. In both multipolar and bipolar neurons in the PNS, Para is present in a proximal region of the axon, comparable to the axonal initial segment (AIS) in vertebrates, 40–60 μm from the soma in multipolar neurons and 20–40 μm in bipolar neurons. Whole-cell reduction ofparaexpression using RNAi in CNs of the adult Johnston’s organ (JO) severely affects sound-evoked potentials (SEPs). However, the duality of Para localization in the CN dendrites and axons identifies a need to develop resources to study compartment-specific roles of proteins that will enable us to better understand Para’s role in mechanosensitive transduction.

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  5. ABSTRACT Hemolymph is driven through the antennae of Drosophila melanogaster by the rhythmic contraction of muscle 16 (m16), which runs through the brain. Contraction of m16 results in the expansion of an elastic ampulla, opening ostia and filling the ampulla. Relaxation of the ampullary membrane forces hemolymph through vessels into the antennae. We show that m16 is an auto-active rhythmic somatic muscle. The activity of m16 leads to the rapid perfusion of the antenna by hemolymph. In addition, it leads to the rhythmic agitation of the brain, which could be important for clearing the interstitial space. 
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  6. Dutzler, Raimund (Ed.)
    Potassium ion (K + ) plays a critical role as an essential electrolyte in all biological systems. Genetically-encoded fluorescent K + biosensors are promising tools to further improve our understanding of K + -dependent processes under normal and pathological conditions. Here, we report the crystal structure of a previously reported genetically-encoded fluorescent K + biosensor, GINKO1, in the K + -bound state. Using structure-guided optimization and directed evolution, we have engineered an improved K + biosensor, designated GINKO2, with higher sensitivity and specificity. We have demonstrated the utility of GINKO2 for in vivo detection and imaging of K + dynamics in multiple model organisms, including bacteria, plants, and mice. 
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