The gene records representing phenol oxidase genes in FlyBase had a number of errors prior to release FB_2009_09, including genes being incorrectly merged together. The gene records have been reassessed for release FB_2009_09 and information has been reassigned to try to correct these errors. The following describes the problems prior to FB_2009_09 and the evidence for splitting the data into a number of separate genes for release FB_2009_09.

The Black cells (Bc) mutation had been merged with the gene corresponding to the CG5779 annotation in FlyBase (this gene encodes the A1 component of phenol oxidase originally described in FBrf0017315) under the single gene record FBgn0000165, despite no study having established that Bc was a mutation of CG5779. The confusion probably arose because phenol oxidase activity is reduced in Bc heterozygous larval cell-free extracts and undetectable in homozygous larval cell-free extracts and because the Bc mutation and the gene corresponding to the CG5779 annotation map to the same location (2-80.6). FBrf0035582 suggests that the primary defect in Bc mutants involves the loss of the physical barrier between phenol oxidase and its substrate in the crystal cells, and that the Bc⁺ product is required for binding the substrate in the paracrystalline structures of the crystal cells so that it is inaccessible to the phenol oxidase enzyme (rather than Bc directly encoding a phenol oxidase). The Bc mutant locus and the gene corresponding to the CG5779 annotation have thus been split into two separate genes in FlyBase in release FB_2009_09. The gene corresponding to the CG5779 annotation has been called both "Dox-A1, Diphenol oxidase A1 subunit" and "Mox, Monophenol oxidase" in the literature. The gene product has been shown to have both monophenol oxidase and diphenol oxidase activity (FBrf0058767). To avoid confusion, the name used for this gene in FlyBase is now "proPO-A1, prophenol oxidase A1" to indicate that the CG5779 annotation encodes the structural locus of the A1 component of phenol oxidase originally described in FBrf0017315, which is produced as a proenzyme. The symbol of the annotation has also been renamed from CG5779 to CG42639 in release 5.22 of the genome annotation to try to avoid confusion. In addition, the "Phox, Phenol oxidase" gene originally described in FBrf0034818 had been merged with the proPO-A1 gene (and thus with the Black cells (Bc) mutation) in FlyBase. This is probably because in FBrf0064348 it is suggested that Phox may be the same locus as proPO-A1, since it maps to the same location and both proPO-A1 and Phox show monophenol oxidase activity. However, FBrf0036109 determined the molecular weight of Phox to be 54kD, which differs from the weight reported for proPO-A1 in FBrf0027149. Thus the "Phox, Phenol oxidase" gene originally described in FBrf0034818 has also been split out into a separate gene in its own right in FlyBase in release FB_2009_09.

FBrf0044478 showed that lesions in the gene corresponding to the CG10484 annotation reduce the activity of the A2 component of phenol oxidase originally described in FBrf0017315, while the A1 and A3 phenol oxidase components originally described in FBrf0017315 are unaffected in these mutant alleles. Thus FBrf0044478 states that the mutants probably identify a structural locus for the A2 phenol oxidase component and designated this locus "Diphenol oxidase-A2, Dox-A2". However, cloning of the gene in FBrf0054017 revealed that the encoded protein had no homology to non-insect phenol oxidases at that time, but instead shows high amino acid identity (57% over the entire length of the D. melanogaster protein) to the mouse Psmd3 gene (accession number MGI:48687), which encodes a proteasome subunit. FBrf0128494 showed that the protein encoded by the CG10484 annotation is a subunit of the regulatory complex of the 26S proteasome in D. melanogaster. FBrf0058767 suggests that the A2 component of phenol oxidase seen in FBrf0017315 could have been an artificial derivative produced from either the A1 or A3 components by the activity of endogenous proteases during extraction, since protease inhibitors were not used in sample preparation (the A2 phenol oxidase component was not detected in FBrf0058767 where serine proteases inhibitors were present during sample preparation). Alternatively, FBrf0027144 shows that the speck (sp) mutants have a marked decreased in the amount of the A2 component of phenol oxidase, and thus FBrf0044480 notes that the A2 component may be a dimer of two non-identical subunits coded for by "Dox-A2" and sp. Given the uncertainty over whether the gene corresponding to the CG10484 annotation does encode a structural locus for the A2 component of phenol oxidase, but the good evidence that it is a proteasome subunit, the gene has been renamed to "Rpn3, Regulatory particle non-ATPase 3" following the nomenclature used in FBrf0152082, which uses the nomenclature used for S. cerevisiae proteasome regulatory particle subunits described in PubMed:9697412. In addition, the annotation corresponding to the gene has been renamed from CG10484 to CG42641 in release 5.22 of the genome annotation.

The "Dox-3, Dopa oxidase-3" locus originally described in FBrf0043263 had incorrectly been merged with the gene corresponding to the CG2952 annotation in FlyBase, based on the GenBank accession AB055857. The merged gene was called "Dox-A3, Diphenol oxidase A3" (FBgn0000487) in FlyBase, since it had been proposed (in FBrf0043263) that the Dox-3 locus was the structural locus for the A3 phenol oxidase component originally described in FBrf0017315. However, as pointed out in FBrf0207791, the AB055857 accession is chimeric, containing portions of CG2952 and CG8193 (both of are predicted to encode prophenol oxidases based on their sequence) and also of CG14196. Evidence in FBrf0207791 (developmental expression profile, immunoblot analyses) indicates that CG2952 does not encode the A3 phenol oxidase component originally described in FBrf0017315 and it is more likely that CG8193 encodes the A3 component. However FBrf0207791 notes there is a discrepancy between the location of CG8193 and that of the mutant Dox-3 strains described in FBrf0043263. Thus it is possible that these mutant strains do not affect the structural locus for the A3 phenol oxidase component originally described in FBrf0017315, but instead represent a modifying factor. Due to the uncertainty over the various components, the "Dox-A3" (FBgn0000487) gene has been split into two separate genes in FlyBase in release FB_2009_09: a gene representing the CG2952 annotation (this annotation and the gene symbol have been renamed to CG42640 in release 5.22 of the genome annotation to avoid confusion) and "Dox-3, Dopa oxidase-3", representing the locus originally described in FBrf0043263. The CG8193 gene record (FBgn0033367) remains as a separate gene record in FlyBase until more evidence is available to determine whether or not it encodes the structural locus for the A3 phenol oxidase and whether or not the Dox-3 strains affect this locus.

The list of genes resulting from this correction of data and their symbols in release FB_2009_09 is:

Symbol: Bc

Name: Black cells

FlyBase ID: FBgn0261382

Annotation symbol: none - not yet cloned

Symbol: proPO-A1

Name: prophenol oxidase A1

FlyBase ID: FBgn0261362

Annotation symbol: CG42639

Comment: Encodes the A1 phenol oxidase component originally described in FBrf0017315.

Symbol: Phox

Name: Phenol oxidase

FlyBase ID: FBgn0261374

Annotation symbol: none - not yet cloned

Symbol: Rpn3

Name: Regulatory particle non-ATPase 3

FlyBase ID: FBgn0261364

Annotation symbol: CG42641

Symbol: CG42640

Name: none

FlyBase ID: FBgn0261363

Annotation symbol: CG42640

Comment: Does not encode the A3 phenol oxidase component originally described in FBrf0017315.

Symbol: Dox-3

Name: Dopa oxidase-3

FlyBase ID: FBgn0261375

Annotation symbol: none - not yet cloned

Comment: May affect the A3 phenol oxidase component originally described in FBrf0017315.

Symbol: CG8193

Name: none

FlyBase ID: FBgn0033367

Annotation symbol: CG8193

Comment: Probably encodes the A3 phenol oxidase component originally described in FBrf0017315.