Gene pink and its linkage to the biogenesis of lysosome organelle complex-2 (BLOC-2)

Introduction
Mutation is defined as the changing of a gene to a variant form; changes which can lead
to discrepancies that can be physically seen. Sometimes these phenotypic changes are the sole
result of a specific genes alteration, but there is always the possibility that the expression of the
original gene has been transformed in such a way that there is an innate change in the organism
in congruence with the phenotypic divergence. In other words, it is common for a physical
variation to serve as an indicator for an underlying gene expression; something not possible to be
seen with the naked human eye. The organism most commonly used to study mutations and the
subsequent effects on an organisms behavior and lifestyle is the Drosophila melanogaster, more
universally known as the common fruit fly. The D. melanogaster exists as an ideal model
organism due to its fulfillment of several important conditions: an efficient reproduction cycle,
affordability, and a short life span. The one flaw of the D. melanogaster existed in the diminutive
size of its chromosomes; the size making it hard to identify changes to the genetic code. This was
solved through the discovery of a certain genre of chromosomes on the salivary gland; more
specifically, the finding of the polytene chromosomes. These chromosomes were developed into
cytogenetic maps and used to offset the flys weakness (Sang 2001). With its one major flaw
accounted for, the Drosophila melanogaster was such an ideal candidate for analysis that it
became fervently used and resulted in it becoming one of the few organisms whose entire
genome is known.
The mutation studied will be explicitly related to the eye color of the D. melanogaster.
Compelling evidence exists that the organelles containing pigments are actually endosomes or
lysosomes. This suggests that mutations affecting the delivery of proteins to the lysosomes

would also affect eye color (Lloyd et al. 1998). Ommochromes and pteridines are what give the
Drosophila the red color of its compound eyes. These pigments, brown and red respectively, fill
the granules which make up the pigment cells; these, in turn, make up the 800 individual
elements of the ommatidia (Lloyd et al. 1998). Each of these pigments has their own respective
biosynthetic pathway. To arrive at the brown pigment, Xanthommatin, found in the fruit fly, there
are several oxidation steps that must take place prior each accelerated by its own enzyme.
These steps are shown below in figure 1 where A, B, C, and D are enzymes tryptophan
oxygenase, kynurenine formamidase, kynurenine hydroxylase, and phenoxazinone synthase
respectively (Summers et al, 1982).

Each of these enzymes are responsible for a different shade of the final resulting pigment
Xanthommatin; therefore by mutating a specific enzyme, a deficiency in Xanthommatin resulting
in the creation of a new pigment. The same theoretical aspect applies to the other pigment,
pteridines, pathway. There are multiple major derivatives of the group pteridine found in the
melanogaster, but the pathway shown in figure 2 relates only to that relevant to drosopterin with
enzymes A and B being GTP cyclohydrolase and drosopterin synthase respectively (Summers et
al. 1982).

Mutation can occur in one or both of these pathways; the resulting eye-color mutation depending
on the locus or loci affected.

The gene pink is an alteration in the normally vivid red color to that of a duller brick
color and is created from the mutation of both the pteridine and ommochrome biosynthetic
pathways. Through test crossing, Morgan discovered the pink mutation directly after the
discovery of his infamous and first mutation, white. A gene allelic to p was first isolated as
CG9770 through the hypothesis that BLOC subunits might have considerable sway over the
biochemical pathways of pigments in the eye (Falcon-Perez et al. 2007). The location was
confirmed through the use of a pink-EGFP plasmid vector in P-element insertion in conjunction
with the Southern blot analysis of radiation (Syrzycka et al. 2007). The relation of pink to the
BLOC-2 subunit resulted in a significant breakthrough in a human genetic disorder.
Phenotypic Characteristics of Mutations in the Gene
The pink gene is located on the third chromosome, right arm, and at position 48.0. Its
characterized based on its desaturation of the normal red color found in the fruit flies (Silva
1988). Morgan discovered that this mutation was recessive and that its mode of inheritance was
not linked to sex. Through it having no linkage to second chromosome genes, Morgan and
Bridges defined it to be autosomal and on the third chromosome. As a recessive gene, the gene
will only be expressed homozygous. Doubly homozygous p, pp, or psnb does not result in
lethality; instead, there is a more extreme form of the mutation expressed. In other words, there
is a marked decrease in the original red pigmentation; enough so that it is possible to easily
distinguish a single mutation color (Falcon-Perez et al. 2007). When carmine (car) and pp were
generated as a doubly homozygous mutant, results were observed showing that incomplete
dominance exists between the two. The resultant phenotype was a mixture between the highest
extremes of car and pp (Falcon-Perez et al. 2007).

One allele of the gene, sunburst (psnb), (its color differing only slightly from pink in that it
was more of a chestnut color) was classified through the observance of the polytence
chromosomes (Silva 1988). Another mutant allele, peach (pp), also results in a decreased
appearance of pigmentation, for this particular allele to that of a more orange color. A third
mutation of the pink gene would be CG9770, a Drosophila orthologue (Falcon-Perez et al.
2007).
Molecular Characteristics of the Gene and the Gene Product
Epistasis was not seen between specific alleles of the pink gene but between p and car as
previously mentioned. The car mutation affects the transport of white and brown proteins while
p affects that of both pathways. As a result of using car and pp as homozygous double mutants,
the brown pigment would be lower than car whereas the red pigmentation would be higher than
that of car (Falcon-Perez et al. 2007).
The two alleles of pink previously mentioned were psnb and pp; their relation to p is
through a frameshift and silent mutation in their coding. The observed mutation is caused only
by a single base pair deletion (Syrzycka et al. 2007) as a silent mutation does not result in change
of the expressed function. It is believed that the alleles marked decrease in pigmentation is due
to the White protein ABC8s decrease in the transport of proteins (Syrzycka et al. 2007).
In addition, pinks function has not been addressed, in literature, at the molecular level.
There are, however, other aspects to consider. The Pink protein is predicted to be eight hundred
and twenty six amino acids long with its allelic gene product, CG9770-PA (from the gene
CG9770), found to have similar amino acid sequence (approximately 22%) to that of protein
complex BLOC-2 (Syrzycka et al. 2007). This gene found in the D. melanogaster is an ortholog

to HPS5 human gene which is responsible for the genetic disorder known as Hermansky-Pudlak
syndrome (HPS) and has been used synonymously with pink (Falcon-Perez et al. 2007).
Conclusion
The discovery of CG9770 Drosophila genes relation to that of the human gene HPS5h
shows the existence of a relationship between the eye color in flies and lysosome biogenesis. As
mentioned, the CG9770 (located on pinks locus) and HPS5h genes are related to the BLOC-2
complex. The BLOC-2 (Biogenesis of Lysosome-related Organelles Complex-2) complex is
believed to be directly correlated to HPS protein HPS5 where HPS is an albinism disorder
marked by excessive and protracted bleeding (Di Pietro et al. 2004). The pink gene is important
in that of what the future could hold for it; CG9770 marks a huge leap of progress for those with
HPS. In order for further progress to be made, however, the function of HPS5, HPS3, and HPS6
should be studied and documented as they are not yet known (Syrzycka et al. 2007).
Evolutionary relationships were explored by comparing homologs of the Drosophila
HPS5 through that of other species genomes including humans, chicken, mice and beetles. As a
result of the evolutionary trees analysis, existence of proteins like CG9770-PA and HPS5 were
found in the studied species; for example, a paralog of HPS5 was found in the human genome
known as LOC9895. Through the order of divergence, it became an accepted theory that the
human HPS5 originated from CG9770 by order of a single gene duplication (Falcon-Perez et al.
2007).
In addition to the suggestion previously mentioned, pinks molecular function should be
delved into, studied, found and recorded. As it is now, all that is known about pink is in its
superficial, phenotypic qualities and in its relationship to CG9770. It stands to reason that
figuring out the molecular function of pink would further research on HPS5 and in its

relationship to lysozyme biogenesis in that of the eyes of the Drosophila melanogaster. These
suggestions seem to be where research into pink is headed as the genes most relevant function
seems to be directly correlated with HPS5.
References
1. Falcn-Prez J. M., R. Romero-Caldern, E.S. Brooks, D.E. Krantz , E.C. Dell'Angelica,
2007 The Drosophila pigmentation gene pink (p) encodes a homolog of human
HermanskyPudlak syndrome 5 (HPS5). Traffic 8: 154-158.
2. Syrzycka M., L. A. McEachern, J. Kinneard, K. Prabhu, Fitzpatrick K., et al., 2007 The
pink gene encodes the Drosophila orthologue of the human HermanskyPudlak syndrome
5 (HPS5) gene. Genome 50: 548-556.
3. Di Pietro S. M., J.M. Falcon-Perez, E.C. Dell'Angelica, 2004 Characterization of BLOC2, a Complex Containing the HermanskyPudlak Syndrome Proteins HPS3, HPS5 and
HPS6. Traffic 5: 276283.
4. Lloyd V., M. Ramaswami, H. Kramer, 1998 Not just pretty eyes: Drosophila eye-colour
mutations and lysosomal delivery. Trends Cell Biology 8: 257-259.
5. Silva F. J., 1988 pink and sunburst mutations of Drosophila melanogaster are alleles of
the same locus. D. I. S. 67: 72.
6. Sang J. H., 2001 Drosophila Melanogaster: The Fruit Fly. Encyclopedia of Genetics:
157-160.
7. Summers K. M., A. J. Howells, N.A. Pyliotis, 1982 Biology of eye pigmentation in
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