Limits...
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.

Show MeSH

Related in: MedlinePlus

Drosophila CG12120 Is Conserved across Insects and Is Related to Fungal IAT(A) Alignment of D. melanogaster, D. pseudoobscura, and A. gambiae CG12120 orthologs and P. chrysogenum IAT protein. Residues in blue are identical (*) across all four species, and residues in green are functionally similar, showing strong (:) or weak (.) similarity. Residue highlighted “1” indicates conserved arginine residue at position 217 that is mutated to proline in tan1 and tan4 mutants. Residue highlighted “2” indicates methionine residue at position 256 that is mutated to isoleucine in tan5 mutant. Cyan rectangle indicates auto-processing site of P. chrysogenum at which pro-IAT is cleaved between glycine 102 and cysteine 103.(B) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) among Drosophila spp., A. gambiae, and P. chrysogenum proteins.(C) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) of N-terminal Drosophila spp., A. gambiae, and P. chrysogenum proteins with presumptive B. mori CG12120 ortholog (see text).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC1285064&req=5

pgen-0010063-g003: Drosophila CG12120 Is Conserved across Insects and Is Related to Fungal IAT(A) Alignment of D. melanogaster, D. pseudoobscura, and A. gambiae CG12120 orthologs and P. chrysogenum IAT protein. Residues in blue are identical (*) across all four species, and residues in green are functionally similar, showing strong (:) or weak (.) similarity. Residue highlighted “1” indicates conserved arginine residue at position 217 that is mutated to proline in tan1 and tan4 mutants. Residue highlighted “2” indicates methionine residue at position 256 that is mutated to isoleucine in tan5 mutant. Cyan rectangle indicates auto-processing site of P. chrysogenum at which pro-IAT is cleaved between glycine 102 and cysteine 103.(B) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) among Drosophila spp., A. gambiae, and P. chrysogenum proteins.(C) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) of N-terminal Drosophila spp., A. gambiae, and P. chrysogenum proteins with presumptive B. mori CG12120 ortholog (see text).

Mentions: CG12120 encodes a 387-amino-acid polypeptide with a predicted molecular weight of approximately 44 kD. CG12120 homologs are present in all sequenced Drosophila genomes, as well as in the genomes of Anopheles gambiae and Bombyx mori. Sequence identity among insect CG12120 proteins extends over the entire length of the protein for the two Diptera (79.8% identical between D. melanogaster and A. gambiae;Figure 3A and 3B) and the first 230–240 amino acids between dipteran and Bombyx mori sequences (45%–48% identical between dipterans and B. mori;Figure 3C). The B. mori sequence may be an incomplete fragment, to be clarified pending the release of an annotated version of the genome sequence. Interestingly, several fungal IATs (Pencillium chrysogenum shown in Figure 3A and 3B) are approximately 50% similar (based on Gonnet series in CLUSTALW [21]) and 20% identical to insect CG12120 proteins, suggesting the conservation of a very ancient gene present in the common ancestor of fungi and metazoans. Fungal IATs are one of three enzymes in the penicillin biosynthetic pathway and catalyze the substitution of the L-α-aminoadipyl side chain of isopenicillin-N with aromatic acyl side chains [22]. A BLAST search of P. chrysogenum IAT to the D. melanogaster genome turned up two other proteins, but these have less substantial similarity to IAT: CG12140, a predicted electron-transferring flavoprotein dehydrogenase (25% identical, 38% similar over a 148-amino-acid region from residues 90–228 of IAT), and CG8864, a predicted monooxygenase/oxidoreductase electron transporter (29% identical, 38% similar over a 78-amino-acid region from residues 224–289 of IAT). Therefore, it appears that CG12120 is the only protein in the Drosophila genome with strong homology to fungal IATs.


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)

Drosophila CG12120 Is Conserved across Insects and Is Related to Fungal IAT(A) Alignment of D. melanogaster, D. pseudoobscura, and A. gambiae CG12120 orthologs and P. chrysogenum IAT protein. Residues in blue are identical (*) across all four species, and residues in green are functionally similar, showing strong (:) or weak (.) similarity. Residue highlighted “1” indicates conserved arginine residue at position 217 that is mutated to proline in tan1 and tan4 mutants. Residue highlighted “2” indicates methionine residue at position 256 that is mutated to isoleucine in tan5 mutant. Cyan rectangle indicates auto-processing site of P. chrysogenum at which pro-IAT is cleaved between glycine 102 and cysteine 103.(B) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) among Drosophila spp., A. gambiae, and P. chrysogenum proteins.(C) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) of N-terminal Drosophila spp., A. gambiae, and P. chrysogenum proteins with presumptive B. mori CG12120 ortholog (see text).
© Copyright Policy
Related In: Results  -  Collection

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

pgen-0010063-g003: Drosophila CG12120 Is Conserved across Insects and Is Related to Fungal IAT(A) Alignment of D. melanogaster, D. pseudoobscura, and A. gambiae CG12120 orthologs and P. chrysogenum IAT protein. Residues in blue are identical (*) across all four species, and residues in green are functionally similar, showing strong (:) or weak (.) similarity. Residue highlighted “1” indicates conserved arginine residue at position 217 that is mutated to proline in tan1 and tan4 mutants. Residue highlighted “2” indicates methionine residue at position 256 that is mutated to isoleucine in tan5 mutant. Cyan rectangle indicates auto-processing site of P. chrysogenum at which pro-IAT is cleaved between glycine 102 and cysteine 103.(B) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) among Drosophila spp., A. gambiae, and P. chrysogenum proteins.(C) Percent sequence identity (blue in upper right) and sequence similarity (red in lower left) of N-terminal Drosophila spp., A. gambiae, and P. chrysogenum proteins with presumptive B. mori CG12120 ortholog (see text).
Mentions: CG12120 encodes a 387-amino-acid polypeptide with a predicted molecular weight of approximately 44 kD. CG12120 homologs are present in all sequenced Drosophila genomes, as well as in the genomes of Anopheles gambiae and Bombyx mori. Sequence identity among insect CG12120 proteins extends over the entire length of the protein for the two Diptera (79.8% identical between D. melanogaster and A. gambiae;Figure 3A and 3B) and the first 230–240 amino acids between dipteran and Bombyx mori sequences (45%–48% identical between dipterans and B. mori;Figure 3C). The B. mori sequence may be an incomplete fragment, to be clarified pending the release of an annotated version of the genome sequence. Interestingly, several fungal IATs (Pencillium chrysogenum shown in Figure 3A and 3B) are approximately 50% similar (based on Gonnet series in CLUSTALW [21]) and 20% identical to insect CG12120 proteins, suggesting the conservation of a very ancient gene present in the common ancestor of fungi and metazoans. Fungal IATs are one of three enzymes in the penicillin biosynthetic pathway and catalyze the substitution of the L-α-aminoadipyl side chain of isopenicillin-N with aromatic acyl side chains [22]. A BLAST search of P. chrysogenum IAT to the D. melanogaster genome turned up two other proteins, but these have less substantial similarity to IAT: CG12140, a predicted electron-transferring flavoprotein dehydrogenase (25% identical, 38% similar over a 148-amino-acid region from residues 90–228 of IAT), and CG8864, a predicted monooxygenase/oxidoreductase electron transporter (29% identical, 38% similar over a 78-amino-acid region from residues 224–289 of IAT). Therefore, it appears that CG12120 is the only protein in the Drosophila genome with strong homology to fungal IATs.

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