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Chromatin Modulatory Proteins and Olfactory Receptor Signaling in the Refinement and Maintenance of Fruitless Expression in Olfactory Receptor Neurons.

Hueston CE, Olsen D, Li Q, Okuwa S, Peng B, Wu J, Volkan PC - PLoS Biol. (2016)

Bottom Line: This regulation requires the chromatin modulatory protein Alhambra (Alh).Our results highlight two feed-forward regulatory mechanisms with both developmentally hardwired and olfactory receptor activity-dependent components that establish and maintain fru expression in ORNs.Such a dual mechanism of fru regulation in ORNs might be a trait of neurons driving plastic aspects of sex-specific behaviors.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, Duke University, Durham, North Carolina, United States of America.

ABSTRACT
During development, sensory neurons must choose identities that allow them to detect specific signals and connect with appropriate target neurons. Ultimately, these sensory neurons will successfully integrate into appropriate neural circuits to generate defined motor outputs, or behavior. This integration requires a developmental coordination between the identity of the neuron and the identity of the circuit. The mechanisms that underlie this coordination are currently unknown. Here, we describe two modes of regulation that coordinate the sensory identities of Drosophila melanogaster olfactory receptor neurons (ORNs) involved in sex-specific behaviors with the sex-specific behavioral circuit identity marker fruitless (fru). The first mode involves a developmental program that coordinately restricts to appropriate ORNs the expression of fru and two olfactory receptors (Or47b and Ir84a) involved in sex-specific behaviors. This regulation requires the chromatin modulatory protein Alhambra (Alh). The second mode relies on the signaling from the olfactory receptors through CamK and histone acetyl transferase p300/CBP to maintain ORN-specific fru expression. Our results highlight two feed-forward regulatory mechanisms with both developmentally hardwired and olfactory receptor activity-dependent components that establish and maintain fru expression in ORNs. Such a dual mechanism of fru regulation in ORNs might be a trait of neurons driving plastic aspects of sex-specific behaviors.

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fru-positive OR expression in at4 sensilla expands to developmentally related fru-negative ORNs in alh mutants. A) Adult antennae and brains labeled with Or47bGal4 UAS-CD8GFP (green) in wild type and alh mutant clones. Magenta staining in brains is against N-cadherin, a neuropil marker. B) Adult antennae and brains labeled with Or88aGal4 UAS-CD8GFP in wild-type and alh mutant clones. C) Adult antennae and brains labeled with Or65aGal4 UAS-CD8GFP in wild-type and alh mutant clones. D–G) Quantification of cell bodies observed in the adult antennae of WT and alh mutant clones. For all graphs, asterisks indicate significant (p < .05) differences from wild type. Error bars represent standard error of the mean (SEM). An ANOVA was performed for each cell type and followed with Tukey’s Honest Significant Difference (HSD)—see Materials and Methods. D) Total Or47b-positive cells. Wild type flies were significantly different from both alh conditions (p < .0001). n = 10–40. All count data may be found in the Supporting Information as S1 Data. E) Total Or88a-positive cells. Both alh conditions were significantly different from wild-type males (p < .0001). n = 27 − 57. F) Total Or65a-positive cells. Both alh conditions were significantly different from wild-type males (p < .05). n = 9–27. G) Total or47b-positive clusters, normalized by total Or47b-positive cells. Wild type flies were significantly different from all alh conditions (p < .0001). n = 10–40. H) Model: in alh mutants, the Or47b odorant receptor expression is expanded to the other ORNs in the at4 sensilla, at the expense of their native OR expression, but the axons of these ORNs continue to target their original locations in the antennal lobe.GENOTYPES:A) eyflp; Or47bGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FB) eyflp; Or88aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FC) eyflp; Or65aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2F
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pbio.1002443.g001: fru-positive OR expression in at4 sensilla expands to developmentally related fru-negative ORNs in alh mutants. A) Adult antennae and brains labeled with Or47bGal4 UAS-CD8GFP (green) in wild type and alh mutant clones. Magenta staining in brains is against N-cadherin, a neuropil marker. B) Adult antennae and brains labeled with Or88aGal4 UAS-CD8GFP in wild-type and alh mutant clones. C) Adult antennae and brains labeled with Or65aGal4 UAS-CD8GFP in wild-type and alh mutant clones. D–G) Quantification of cell bodies observed in the adult antennae of WT and alh mutant clones. For all graphs, asterisks indicate significant (p < .05) differences from wild type. Error bars represent standard error of the mean (SEM). An ANOVA was performed for each cell type and followed with Tukey’s Honest Significant Difference (HSD)—see Materials and Methods. D) Total Or47b-positive cells. Wild type flies were significantly different from both alh conditions (p < .0001). n = 10–40. All count data may be found in the Supporting Information as S1 Data. E) Total Or88a-positive cells. Both alh conditions were significantly different from wild-type males (p < .0001). n = 27 − 57. F) Total Or65a-positive cells. Both alh conditions were significantly different from wild-type males (p < .05). n = 9–27. G) Total or47b-positive clusters, normalized by total Or47b-positive cells. Wild type flies were significantly different from all alh conditions (p < .0001). n = 10–40. H) Model: in alh mutants, the Or47b odorant receptor expression is expanded to the other ORNs in the at4 sensilla, at the expense of their native OR expression, but the axons of these ORNs continue to target their original locations in the antennal lobe.GENOTYPES:A) eyflp; Or47bGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FB) eyflp; Or88aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FC) eyflp; Or65aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2F

Mentions: We previously carried out a forward genetic screen for regulators of class-specific ORN development in the olfactory system [24]. This was a histology-based mutagenesis screen where the glomerular targeting patterns of three different classes of ORNs (Or47a, Or47b, and Gr21a) were analyzed in antennal mutant clones in an otherwise heterozygous animal [25]. We specifically looked for mutants with defective ORN projection patterns in the antennal lobe. In this screen, we isolated a mutation (p1353) that modified the projection pattern of Or47b ORNs in the antennal lobe (Fig 1). We mapped this mutation to the D. melanogaster orthologue of the chromatin modulatory protein AF10, Alhambra (alh). We refer to this mutation as alh1353 in the rest of the paper. In both alh1353 and alhJ8C8 mutants, Or47b ORN axons project to their normal target, the VA1v glomerulus, as well as the DL3 glomerulus (Fig 1A and 1H). In addition, the VA1v glomerulus expands dorsally, appearing to almost engulf the VA1d glomerulus, leading to a loss of VA1d glomerulus in some of the mutant antennal lobes [26]. The DL3 and VA1d glomeruli are normally innervated by other at4 sensilla ORNs that express Or65a and Or88a, respectively, which develop from the same sensory organ precursor as Or47b ORNs [20].


Chromatin Modulatory Proteins and Olfactory Receptor Signaling in the Refinement and Maintenance of Fruitless Expression in Olfactory Receptor Neurons.

Hueston CE, Olsen D, Li Q, Okuwa S, Peng B, Wu J, Volkan PC - PLoS Biol. (2016)

fru-positive OR expression in at4 sensilla expands to developmentally related fru-negative ORNs in alh mutants. A) Adult antennae and brains labeled with Or47bGal4 UAS-CD8GFP (green) in wild type and alh mutant clones. Magenta staining in brains is against N-cadherin, a neuropil marker. B) Adult antennae and brains labeled with Or88aGal4 UAS-CD8GFP in wild-type and alh mutant clones. C) Adult antennae and brains labeled with Or65aGal4 UAS-CD8GFP in wild-type and alh mutant clones. D–G) Quantification of cell bodies observed in the adult antennae of WT and alh mutant clones. For all graphs, asterisks indicate significant (p < .05) differences from wild type. Error bars represent standard error of the mean (SEM). An ANOVA was performed for each cell type and followed with Tukey’s Honest Significant Difference (HSD)—see Materials and Methods. D) Total Or47b-positive cells. Wild type flies were significantly different from both alh conditions (p < .0001). n = 10–40. All count data may be found in the Supporting Information as S1 Data. E) Total Or88a-positive cells. Both alh conditions were significantly different from wild-type males (p < .0001). n = 27 − 57. F) Total Or65a-positive cells. Both alh conditions were significantly different from wild-type males (p < .05). n = 9–27. G) Total or47b-positive clusters, normalized by total Or47b-positive cells. Wild type flies were significantly different from all alh conditions (p < .0001). n = 10–40. H) Model: in alh mutants, the Or47b odorant receptor expression is expanded to the other ORNs in the at4 sensilla, at the expense of their native OR expression, but the axons of these ORNs continue to target their original locations in the antennal lobe.GENOTYPES:A) eyflp; Or47bGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FB) eyflp; Or88aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FC) eyflp; Or65aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2F
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4836687&req=5

pbio.1002443.g001: fru-positive OR expression in at4 sensilla expands to developmentally related fru-negative ORNs in alh mutants. A) Adult antennae and brains labeled with Or47bGal4 UAS-CD8GFP (green) in wild type and alh mutant clones. Magenta staining in brains is against N-cadherin, a neuropil marker. B) Adult antennae and brains labeled with Or88aGal4 UAS-CD8GFP in wild-type and alh mutant clones. C) Adult antennae and brains labeled with Or65aGal4 UAS-CD8GFP in wild-type and alh mutant clones. D–G) Quantification of cell bodies observed in the adult antennae of WT and alh mutant clones. For all graphs, asterisks indicate significant (p < .05) differences from wild type. Error bars represent standard error of the mean (SEM). An ANOVA was performed for each cell type and followed with Tukey’s Honest Significant Difference (HSD)—see Materials and Methods. D) Total Or47b-positive cells. Wild type flies were significantly different from both alh conditions (p < .0001). n = 10–40. All count data may be found in the Supporting Information as S1 Data. E) Total Or88a-positive cells. Both alh conditions were significantly different from wild-type males (p < .0001). n = 27 − 57. F) Total Or65a-positive cells. Both alh conditions were significantly different from wild-type males (p < .05). n = 9–27. G) Total or47b-positive clusters, normalized by total Or47b-positive cells. Wild type flies were significantly different from all alh conditions (p < .0001). n = 10–40. H) Model: in alh mutants, the Or47b odorant receptor expression is expanded to the other ORNs in the at4 sensilla, at the expense of their native OR expression, but the axons of these ORNs continue to target their original locations in the antennal lobe.GENOTYPES:A) eyflp; Or47bGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or47bGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FB) eyflp; Or88aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or88aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2FC) eyflp; Or65aGal4/UAS-CD8GFP; FRT82/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alh1353/FRT82Gal80E2F,eyflp; Or65aGal4/UAS-CD8GFP; FRT82alhj8c8/FRT82Gal80E2F
Mentions: We previously carried out a forward genetic screen for regulators of class-specific ORN development in the olfactory system [24]. This was a histology-based mutagenesis screen where the glomerular targeting patterns of three different classes of ORNs (Or47a, Or47b, and Gr21a) were analyzed in antennal mutant clones in an otherwise heterozygous animal [25]. We specifically looked for mutants with defective ORN projection patterns in the antennal lobe. In this screen, we isolated a mutation (p1353) that modified the projection pattern of Or47b ORNs in the antennal lobe (Fig 1). We mapped this mutation to the D. melanogaster orthologue of the chromatin modulatory protein AF10, Alhambra (alh). We refer to this mutation as alh1353 in the rest of the paper. In both alh1353 and alhJ8C8 mutants, Or47b ORN axons project to their normal target, the VA1v glomerulus, as well as the DL3 glomerulus (Fig 1A and 1H). In addition, the VA1v glomerulus expands dorsally, appearing to almost engulf the VA1d glomerulus, leading to a loss of VA1d glomerulus in some of the mutant antennal lobes [26]. The DL3 and VA1d glomeruli are normally innervated by other at4 sensilla ORNs that express Or65a and Or88a, respectively, which develop from the same sensory organ precursor as Or47b ORNs [20].

Bottom Line: This regulation requires the chromatin modulatory protein Alhambra (Alh).Our results highlight two feed-forward regulatory mechanisms with both developmentally hardwired and olfactory receptor activity-dependent components that establish and maintain fru expression in ORNs.Such a dual mechanism of fru regulation in ORNs might be a trait of neurons driving plastic aspects of sex-specific behaviors.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, Duke University, Durham, North Carolina, United States of America.

ABSTRACT
During development, sensory neurons must choose identities that allow them to detect specific signals and connect with appropriate target neurons. Ultimately, these sensory neurons will successfully integrate into appropriate neural circuits to generate defined motor outputs, or behavior. This integration requires a developmental coordination between the identity of the neuron and the identity of the circuit. The mechanisms that underlie this coordination are currently unknown. Here, we describe two modes of regulation that coordinate the sensory identities of Drosophila melanogaster olfactory receptor neurons (ORNs) involved in sex-specific behaviors with the sex-specific behavioral circuit identity marker fruitless (fru). The first mode involves a developmental program that coordinately restricts to appropriate ORNs the expression of fru and two olfactory receptors (Or47b and Ir84a) involved in sex-specific behaviors. This regulation requires the chromatin modulatory protein Alhambra (Alh). The second mode relies on the signaling from the olfactory receptors through CamK and histone acetyl transferase p300/CBP to maintain ORN-specific fru expression. Our results highlight two feed-forward regulatory mechanisms with both developmentally hardwired and olfactory receptor activity-dependent components that establish and maintain fru expression in ORNs. Such a dual mechanism of fru regulation in ORNs might be a trait of neurons driving plastic aspects of sex-specific behaviors.

Show MeSH
Related in: MedlinePlus