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Diverse Roles of Axonemal Dyneins in Drosophila Auditory Neuron Function and Mechanical Amplification in Hearing.

Karak S, Jacobs JS, Kittelmann M, Spalthoff C, Katana R, Sivan-Loukianova E, Schon MA, Kernan MJ, Eberl DF, Göpfert MC - Sci Rep (2015)

Bottom Line: Null alleles of both dyneins equally abolished electrical auditory neuron responses, yet whereas mutations in Dmdnah3 facilitated mechanical amplification, amplification was abolished by mutations in Dmdnai2.This establishes diverse roles of axonemal dyneins in Drosophila auditory neuron function and links auditory neuron motility to primary cilia and axonemal dyneins.Mutant defects in sperm competition suggest that both dyneins also function in sperm motility.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Neurobiology, University of Göttingen, 37077 Göttingen, Germany.

ABSTRACT
Much like vertebrate hair cells, the chordotonal sensory neurons that mediate hearing in Drosophila are motile and amplify the mechanical input of the ear. Because the neurons bear mechanosensory primary cilia whose microtubule axonemes display dynein arms, we hypothesized that their motility is powered by dyneins. Here, we describe two axonemal dynein proteins that are required for Drosophila auditory neuron function, localize to their primary cilia, and differently contribute to mechanical amplification in hearing. Promoter fusions revealed that the two axonemal dynein genes Dmdnah3 (=CG17150) and Dmdnai2 (=CG6053) are expressed in chordotonal neurons, including the auditory ones in the fly's ear. Null alleles of both dyneins equally abolished electrical auditory neuron responses, yet whereas mutations in Dmdnah3 facilitated mechanical amplification, amplification was abolished by mutations in Dmdnai2. Epistasis analysis revealed that Dmdnah3 acts downstream of Nan-Iav channels in controlling the amplificatory gain. Dmdnai2, in addition to being required for amplification, was essential for outer dynein arms in auditory neuron cilia. This establishes diverse roles of axonemal dyneins in Drosophila auditory neuron function and links auditory neuron motility to primary cilia and axonemal dyneins. Mutant defects in sperm competition suggest that both dyneins also function in sperm motility.

No MeSH data available.


Related in: MedlinePlus

Dmdnah3 and Dmdnai2 are expressed in sperm and impair sperm competition.(A) Regulatory sequences of Dmdnah3 (left) and Dmdnai2 (right) fused to Gal4 can both drive expression of fluorescent markers in sperm. Expression is seen both in developing sperm tails in the testis (t) as well as in mature sperm in the seminal vesicles (sv). (B) Mutations in Dmdnah3 and Dmdnai2 impair sperm competition. Dmdnah31and Dmdnai21 mutant males (black bars) and their heterozygous controls (white bars) were tested for their ability to displace sperm in previously mated females. Sperm displacement was measured as the proportion of offspring from the second male (P2/P1 + P2). Bars represent means, error bars represent SEM. Number of males measured for each genotype is shown in or above the bar. **p < 0.01; ****p < 0.0001 (Kruskal-Wallis test followed by two-tailed Mann-Whitney U tests). Data were included only if at least one offspring from the second male was recovered. This conservative approach was to eliminate any possible effect of the mutations on the ability of males to achieve copulation. The numbers of males that were thus not taken into consideration were 0 for Dmdnah31/TM3 but 23 for Dmdnah31/Df, and 1 for Dmdnai21/TM6 male but 4 for Dmdnai21/Dmdnai21.
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f4: Dmdnah3 and Dmdnai2 are expressed in sperm and impair sperm competition.(A) Regulatory sequences of Dmdnah3 (left) and Dmdnai2 (right) fused to Gal4 can both drive expression of fluorescent markers in sperm. Expression is seen both in developing sperm tails in the testis (t) as well as in mature sperm in the seminal vesicles (sv). (B) Mutations in Dmdnah3 and Dmdnai2 impair sperm competition. Dmdnah31and Dmdnai21 mutant males (black bars) and their heterozygous controls (white bars) were tested for their ability to displace sperm in previously mated females. Sperm displacement was measured as the proportion of offspring from the second male (P2/P1 + P2). Bars represent means, error bars represent SEM. Number of males measured for each genotype is shown in or above the bar. **p < 0.01; ****p < 0.0001 (Kruskal-Wallis test followed by two-tailed Mann-Whitney U tests). Data were included only if at least one offspring from the second male was recovered. This conservative approach was to eliminate any possible effect of the mutations on the ability of males to achieve copulation. The numbers of males that were thus not taken into consideration were 0 for Dmdnah31/TM3 but 23 for Dmdnah31/Df, and 1 for Dmdnai21/TM6 male but 4 for Dmdnai21/Dmdnai21.

Mentions: In Drosophila, sperm and auditory chordotonal neurons are the only ciliated cells whose cilia generate motility with axonemal dynein arms. Driving UAS-EGFP via Dmdnah3-GAL4 or Dmdnai2-GAL4 revealed that both Dmdnah3 and Dmdnai2 are expressed in sperm (Fig. 4A), corroborating the reported presence of DmDNAH3 in sperm revealed by mass spectrometry38. We thus analyzed male fertility in Dmdnah31 and Dmdnai21 mutants. Homozygous Dmdnah31 mutants were behaviorally able to mate, but no offspring were recovered, and testis squashes consistently showed sperm that were not motile. Homozygous Dmdnai21 males, by contrast, if they achieved mating, produced as many males as heterozygous controls, and testis squashes from all males showed motile sperm. Offspring was also obtained from Dmdnah31 mutants when the Dmdnah31 mutation was uncovered by the deficiency Df(3L)BSC371, suggesting that the loss of male fertility and sperm motility in the homozygous mutants –but not their hearing defects (Fig. S7)—arise from a secondary mutation. To characterize contributions of DmDNAH3 and DmDNAI2 to the function of sperm, we performed sperm competition assays in which w1118 females were first mated to w; FRT40A, neoFRTG13, w+ males (P1) and then to balanced Dmdnah31/TM3 or Dmdnai21/TM6 flies and mutant Dmdnah31/Df(3L)BSC371 or Dmdnai21/Dmdnai21 males (P2). Scoring the offspring revealed that sperm from control males displaced 80–90% of P1 sperm, but sperm from Dmdnah31/Df(3L)BSC371 males were almost completely unable to compete, and Dmdnah21 homozygotes showed a highly significant reduction in sperm competition (Fig. 4B). This extends the roles of axonemal dyneins in sperm competition39 to DmDNAH3 and DmDNAI2 and shows that these two proteins, in addition to their auditory requirements, contribute to sperm function and motility.


Diverse Roles of Axonemal Dyneins in Drosophila Auditory Neuron Function and Mechanical Amplification in Hearing.

Karak S, Jacobs JS, Kittelmann M, Spalthoff C, Katana R, Sivan-Loukianova E, Schon MA, Kernan MJ, Eberl DF, Göpfert MC - Sci Rep (2015)

Dmdnah3 and Dmdnai2 are expressed in sperm and impair sperm competition.(A) Regulatory sequences of Dmdnah3 (left) and Dmdnai2 (right) fused to Gal4 can both drive expression of fluorescent markers in sperm. Expression is seen both in developing sperm tails in the testis (t) as well as in mature sperm in the seminal vesicles (sv). (B) Mutations in Dmdnah3 and Dmdnai2 impair sperm competition. Dmdnah31and Dmdnai21 mutant males (black bars) and their heterozygous controls (white bars) were tested for their ability to displace sperm in previously mated females. Sperm displacement was measured as the proportion of offspring from the second male (P2/P1 + P2). Bars represent means, error bars represent SEM. Number of males measured for each genotype is shown in or above the bar. **p < 0.01; ****p < 0.0001 (Kruskal-Wallis test followed by two-tailed Mann-Whitney U tests). Data were included only if at least one offspring from the second male was recovered. This conservative approach was to eliminate any possible effect of the mutations on the ability of males to achieve copulation. The numbers of males that were thus not taken into consideration were 0 for Dmdnah31/TM3 but 23 for Dmdnah31/Df, and 1 for Dmdnai21/TM6 male but 4 for Dmdnai21/Dmdnai21.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4660584&req=5

f4: Dmdnah3 and Dmdnai2 are expressed in sperm and impair sperm competition.(A) Regulatory sequences of Dmdnah3 (left) and Dmdnai2 (right) fused to Gal4 can both drive expression of fluorescent markers in sperm. Expression is seen both in developing sperm tails in the testis (t) as well as in mature sperm in the seminal vesicles (sv). (B) Mutations in Dmdnah3 and Dmdnai2 impair sperm competition. Dmdnah31and Dmdnai21 mutant males (black bars) and their heterozygous controls (white bars) were tested for their ability to displace sperm in previously mated females. Sperm displacement was measured as the proportion of offspring from the second male (P2/P1 + P2). Bars represent means, error bars represent SEM. Number of males measured for each genotype is shown in or above the bar. **p < 0.01; ****p < 0.0001 (Kruskal-Wallis test followed by two-tailed Mann-Whitney U tests). Data were included only if at least one offspring from the second male was recovered. This conservative approach was to eliminate any possible effect of the mutations on the ability of males to achieve copulation. The numbers of males that were thus not taken into consideration were 0 for Dmdnah31/TM3 but 23 for Dmdnah31/Df, and 1 for Dmdnai21/TM6 male but 4 for Dmdnai21/Dmdnai21.
Mentions: In Drosophila, sperm and auditory chordotonal neurons are the only ciliated cells whose cilia generate motility with axonemal dynein arms. Driving UAS-EGFP via Dmdnah3-GAL4 or Dmdnai2-GAL4 revealed that both Dmdnah3 and Dmdnai2 are expressed in sperm (Fig. 4A), corroborating the reported presence of DmDNAH3 in sperm revealed by mass spectrometry38. We thus analyzed male fertility in Dmdnah31 and Dmdnai21 mutants. Homozygous Dmdnah31 mutants were behaviorally able to mate, but no offspring were recovered, and testis squashes consistently showed sperm that were not motile. Homozygous Dmdnai21 males, by contrast, if they achieved mating, produced as many males as heterozygous controls, and testis squashes from all males showed motile sperm. Offspring was also obtained from Dmdnah31 mutants when the Dmdnah31 mutation was uncovered by the deficiency Df(3L)BSC371, suggesting that the loss of male fertility and sperm motility in the homozygous mutants –but not their hearing defects (Fig. S7)—arise from a secondary mutation. To characterize contributions of DmDNAH3 and DmDNAI2 to the function of sperm, we performed sperm competition assays in which w1118 females were first mated to w; FRT40A, neoFRTG13, w+ males (P1) and then to balanced Dmdnah31/TM3 or Dmdnai21/TM6 flies and mutant Dmdnah31/Df(3L)BSC371 or Dmdnai21/Dmdnai21 males (P2). Scoring the offspring revealed that sperm from control males displaced 80–90% of P1 sperm, but sperm from Dmdnah31/Df(3L)BSC371 males were almost completely unable to compete, and Dmdnah21 homozygotes showed a highly significant reduction in sperm competition (Fig. 4B). This extends the roles of axonemal dyneins in sperm competition39 to DmDNAH3 and DmDNAI2 and shows that these two proteins, in addition to their auditory requirements, contribute to sperm function and motility.

Bottom Line: Null alleles of both dyneins equally abolished electrical auditory neuron responses, yet whereas mutations in Dmdnah3 facilitated mechanical amplification, amplification was abolished by mutations in Dmdnai2.This establishes diverse roles of axonemal dyneins in Drosophila auditory neuron function and links auditory neuron motility to primary cilia and axonemal dyneins.Mutant defects in sperm competition suggest that both dyneins also function in sperm motility.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Neurobiology, University of Göttingen, 37077 Göttingen, Germany.

ABSTRACT
Much like vertebrate hair cells, the chordotonal sensory neurons that mediate hearing in Drosophila are motile and amplify the mechanical input of the ear. Because the neurons bear mechanosensory primary cilia whose microtubule axonemes display dynein arms, we hypothesized that their motility is powered by dyneins. Here, we describe two axonemal dynein proteins that are required for Drosophila auditory neuron function, localize to their primary cilia, and differently contribute to mechanical amplification in hearing. Promoter fusions revealed that the two axonemal dynein genes Dmdnah3 (=CG17150) and Dmdnai2 (=CG6053) are expressed in chordotonal neurons, including the auditory ones in the fly's ear. Null alleles of both dyneins equally abolished electrical auditory neuron responses, yet whereas mutations in Dmdnah3 facilitated mechanical amplification, amplification was abolished by mutations in Dmdnai2. Epistasis analysis revealed that Dmdnah3 acts downstream of Nan-Iav channels in controlling the amplificatory gain. Dmdnai2, in addition to being required for amplification, was essential for outer dynein arms in auditory neuron cilia. This establishes diverse roles of axonemal dyneins in Drosophila auditory neuron function and links auditory neuron motility to primary cilia and axonemal dyneins. Mutant defects in sperm competition suggest that both dyneins also function in sperm motility.

No MeSH data available.


Related in: MedlinePlus