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Drosophila tan encodes a novel hydrolase required in pigmentation and vision.

True JR, Yeh SD, Hovemann BT, Kemme T, Meinertzhagen IA, Edwards TN, Liou SR, Han Q, Li J - PLoS Genet. (2005)

Bottom Line: We characterized two tan-like P-element insertions that failed to complement classical tan mutations.Both P insertions showed abnormally low transcription of the CG12120 mRNA.We conclude that D. melanogaster CG12120 corresponds to tan.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecology and Evolution, State University of New York, Stony Brook, New York, United States of America. jrtrue@life.bio.sunysb.edu

ABSTRACT
Many proteins are used repeatedly in development, but usually the function of the protein is similar in the different contexts. Here we report that the classical Drosophila melanogaster locus tan encodes a novel enzyme required for two very different cellular functions: hydrolysis of N-beta-alanyl dopamine (NBAD) to dopamine during cuticular melanization, and hydrolysis of carcinine to histamine in the metabolism of photoreceptor neurotransmitter. We characterized two tan-like P-element insertions that failed to complement classical tan mutations. Both are inserted in the 5' untranslated region of the previously uncharacterized gene CG12120, a putative homolog of fungal isopenicillin-N N-acyltransferase (EC 2.3.1.164). Both P insertions showed abnormally low transcription of the CG12120 mRNA. Ectopic CG12120 expression rescued tan mutant pigmentation phenotypes and caused the production of striking black melanin patterns. Electroretinogram and head histamine assays indicated that CG12120 is required for hydrolysis of carcinine to histamine, which is required for histaminergic neurotransmission. Recombinant CG12120 protein efficiently hydrolyzed both NBAD to dopamine and carcinine to histamine. We conclude that D. melanogaster CG12120 corresponds to tan. This is, to our knowledge, the first molecular genetic characterization of NBAD hydrolase and carcinine hydrolase activity in any organism and is central to the understanding of pigmentation and photoreceptor function.

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Tan Expression and Function in the Eye and Optic Lobes(A) RNA in situ hybridization with an antisense tan cDNA probe to wild-type adult head sections. Horizontal section cut through the eye and the optic lobes.(B) Sagittal section through the ommatidial array of the eye.(C) Control sense probe does not show significant staining (horizontal section).(D–G) ERG phenotypes of wild-type flies (D) and tan mutants (E–G). (D) CantonS (wild-type) ERG showing normal “on” and “off” transients. Upper trace in each recording indicates duration of light input stimulus (see Materials and Methods). (E) tan2 mutant showing complete absence of “on” and “off” transients. (F) P{d07784} exhibits a normal “on” and “off” transient even though it has a tan-like pigmentation phenotype. (G) P{g1557} lacks “on” and “off” transients.(H and I) Normalized “on” (H) and “off” (I) transients for wild-type, tan2 (t[2]), tan-like P excision lines, and “revertant” P excision lines (see text) expressed as a percentage of the respective sustained negative potential. Insets: Correlation between head histamine content and the normalized magnitude of ERG ‘on’ and ‘off’ transients (see text). Error bars: ± 1 standard error in both dimensions.(J) Head histamine contents for tan-like and revertant P excision lines (see text), relative to control w1118 and tan1w1118 flies. Head histamine increased in all flies that drank 0.5% carcinine in 4% glucose (black bars), relative to controls that drank only 4% glucose (open bars). Revertant and tan-like excision lines are each arranged in rank order. Note that excision line flies that drank only 4% glucose have histamine contents too small to show above the abscissa. Error bars represent ± 1 standard error.La, lamina; Me, medulla; Re, retina. Scale bars = 50 μm.
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pgen-0010063-g002: Tan Expression and Function in the Eye and Optic Lobes(A) RNA in situ hybridization with an antisense tan cDNA probe to wild-type adult head sections. Horizontal section cut through the eye and the optic lobes.(B) Sagittal section through the ommatidial array of the eye.(C) Control sense probe does not show significant staining (horizontal section).(D–G) ERG phenotypes of wild-type flies (D) and tan mutants (E–G). (D) CantonS (wild-type) ERG showing normal “on” and “off” transients. Upper trace in each recording indicates duration of light input stimulus (see Materials and Methods). (E) tan2 mutant showing complete absence of “on” and “off” transients. (F) P{d07784} exhibits a normal “on” and “off” transient even though it has a tan-like pigmentation phenotype. (G) P{g1557} lacks “on” and “off” transients.(H and I) Normalized “on” (H) and “off” (I) transients for wild-type, tan2 (t[2]), tan-like P excision lines, and “revertant” P excision lines (see text) expressed as a percentage of the respective sustained negative potential. Insets: Correlation between head histamine content and the normalized magnitude of ERG ‘on’ and ‘off’ transients (see text). Error bars: ± 1 standard error in both dimensions.(J) Head histamine contents for tan-like and revertant P excision lines (see text), relative to control w1118 and tan1w1118 flies. Head histamine increased in all flies that drank 0.5% carcinine in 4% glucose (black bars), relative to controls that drank only 4% glucose (open bars). Revertant and tan-like excision lines are each arranged in rank order. Note that excision line flies that drank only 4% glucose have histamine contents too small to show above the abscissa. Error bars represent ± 1 standard error.La, lamina; Me, medulla; Re, retina. Scale bars = 50 μm.

Mentions: In order to verify that tan/CG12120 is expressed in the Drosophila visual system we performed in situ hybridization experiments using a digoxigenin-labeled CG12120 cDNA antisense RNA probe (Figure 2). Head cryosections clearly revealed labeling in the retina. In horizontal sections the label appeared in thread-like structures resembling photoreceptor cell expression (Figure 2A). In sections that cut the retina in a sagittal plane, label showed the typical ommatidial structure of the retina (Figure 2B). Light areas resembling the ommatidial cavity were surrounded by dark label constituting a ring-like structure composed of photoreceptor cell somata. This pattern clearly indicates that the photoreceptor cell expression of tan differs from, and is complementary to, the lamina epithelial glial expression of ebony (see [5]).


Drosophila tan encodes a novel hydrolase required in pigmentation and vision.

True JR, Yeh SD, Hovemann BT, Kemme T, Meinertzhagen IA, Edwards TN, Liou SR, Han Q, Li J - PLoS Genet. (2005)

Tan Expression and Function in the Eye and Optic Lobes(A) RNA in situ hybridization with an antisense tan cDNA probe to wild-type adult head sections. Horizontal section cut through the eye and the optic lobes.(B) Sagittal section through the ommatidial array of the eye.(C) Control sense probe does not show significant staining (horizontal section).(D–G) ERG phenotypes of wild-type flies (D) and tan mutants (E–G). (D) CantonS (wild-type) ERG showing normal “on” and “off” transients. Upper trace in each recording indicates duration of light input stimulus (see Materials and Methods). (E) tan2 mutant showing complete absence of “on” and “off” transients. (F) P{d07784} exhibits a normal “on” and “off” transient even though it has a tan-like pigmentation phenotype. (G) P{g1557} lacks “on” and “off” transients.(H and I) Normalized “on” (H) and “off” (I) transients for wild-type, tan2 (t[2]), tan-like P excision lines, and “revertant” P excision lines (see text) expressed as a percentage of the respective sustained negative potential. Insets: Correlation between head histamine content and the normalized magnitude of ERG ‘on’ and ‘off’ transients (see text). Error bars: ± 1 standard error in both dimensions.(J) Head histamine contents for tan-like and revertant P excision lines (see text), relative to control w1118 and tan1w1118 flies. Head histamine increased in all flies that drank 0.5% carcinine in 4% glucose (black bars), relative to controls that drank only 4% glucose (open bars). Revertant and tan-like excision lines are each arranged in rank order. Note that excision line flies that drank only 4% glucose have histamine contents too small to show above the abscissa. Error bars represent ± 1 standard error.La, lamina; Me, medulla; Re, retina. Scale bars = 50 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC1285064&req=5

pgen-0010063-g002: Tan Expression and Function in the Eye and Optic Lobes(A) RNA in situ hybridization with an antisense tan cDNA probe to wild-type adult head sections. Horizontal section cut through the eye and the optic lobes.(B) Sagittal section through the ommatidial array of the eye.(C) Control sense probe does not show significant staining (horizontal section).(D–G) ERG phenotypes of wild-type flies (D) and tan mutants (E–G). (D) CantonS (wild-type) ERG showing normal “on” and “off” transients. Upper trace in each recording indicates duration of light input stimulus (see Materials and Methods). (E) tan2 mutant showing complete absence of “on” and “off” transients. (F) P{d07784} exhibits a normal “on” and “off” transient even though it has a tan-like pigmentation phenotype. (G) P{g1557} lacks “on” and “off” transients.(H and I) Normalized “on” (H) and “off” (I) transients for wild-type, tan2 (t[2]), tan-like P excision lines, and “revertant” P excision lines (see text) expressed as a percentage of the respective sustained negative potential. Insets: Correlation between head histamine content and the normalized magnitude of ERG ‘on’ and ‘off’ transients (see text). Error bars: ± 1 standard error in both dimensions.(J) Head histamine contents for tan-like and revertant P excision lines (see text), relative to control w1118 and tan1w1118 flies. Head histamine increased in all flies that drank 0.5% carcinine in 4% glucose (black bars), relative to controls that drank only 4% glucose (open bars). Revertant and tan-like excision lines are each arranged in rank order. Note that excision line flies that drank only 4% glucose have histamine contents too small to show above the abscissa. Error bars represent ± 1 standard error.La, lamina; Me, medulla; Re, retina. Scale bars = 50 μm.
Mentions: In order to verify that tan/CG12120 is expressed in the Drosophila visual system we performed in situ hybridization experiments using a digoxigenin-labeled CG12120 cDNA antisense RNA probe (Figure 2). Head cryosections clearly revealed labeling in the retina. In horizontal sections the label appeared in thread-like structures resembling photoreceptor cell expression (Figure 2A). In sections that cut the retina in a sagittal plane, label showed the typical ommatidial structure of the retina (Figure 2B). Light areas resembling the ommatidial cavity were surrounded by dark label constituting a ring-like structure composed of photoreceptor cell somata. This pattern clearly indicates that the photoreceptor cell expression of tan differs from, and is complementary to, the lamina epithelial glial expression of ebony (see [5]).

Bottom Line: We characterized two tan-like P-element insertions that failed to complement classical tan mutations.Both P insertions showed abnormally low transcription of the CG12120 mRNA.We conclude that D. melanogaster CG12120 corresponds to tan.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecology and Evolution, State University of New York, Stony Brook, New York, United States of America. jrtrue@life.bio.sunysb.edu

ABSTRACT
Many proteins are used repeatedly in development, but usually the function of the protein is similar in the different contexts. Here we report that the classical Drosophila melanogaster locus tan encodes a novel enzyme required for two very different cellular functions: hydrolysis of N-beta-alanyl dopamine (NBAD) to dopamine during cuticular melanization, and hydrolysis of carcinine to histamine in the metabolism of photoreceptor neurotransmitter. We characterized two tan-like P-element insertions that failed to complement classical tan mutations. Both are inserted in the 5' untranslated region of the previously uncharacterized gene CG12120, a putative homolog of fungal isopenicillin-N N-acyltransferase (EC 2.3.1.164). Both P insertions showed abnormally low transcription of the CG12120 mRNA. Ectopic CG12120 expression rescued tan mutant pigmentation phenotypes and caused the production of striking black melanin patterns. Electroretinogram and head histamine assays indicated that CG12120 is required for hydrolysis of carcinine to histamine, which is required for histaminergic neurotransmission. Recombinant CG12120 protein efficiently hydrolyzed both NBAD to dopamine and carcinine to histamine. We conclude that D. melanogaster CG12120 corresponds to tan. This is, to our knowledge, the first molecular genetic characterization of NBAD hydrolase and carcinine hydrolase activity in any organism and is central to the understanding of pigmentation and photoreceptor function.

Show MeSH
Related in: MedlinePlus