Biology Reference
In-Depth Information
transposon in a GAL4-dependent fashion. Thus, when used in conjunction with
a particular GAL4 line, these transposons might suppress or enhance the disease
model phenotype as a result of insertional inactivation of the genes they reside in
or as a result of overexpression of flanking genes. These potential effects can be
easily distinguished in subsequent studies. This modifier screening approach is
compatible with a variety of phenotypes and could be used to identify modifiers
of behavioral, morphological, or recessive lethal phenotypes. A particularly
useful attribute of this screening method is that the insertion location of all of
the
lines has been determined and thus modifier genes are readily identified.
Use of the
EP
collection in a modifier screening context has proven extremely
valuable in studies to identify modifiers of polyglutamine, PD, and tau pathology
in Drosophila (Fernandez-Funez
EP
, 2000; Shulman and Feany, 2003).
Finally, a number of Drosophila cell lines are available for cell biological
studies of neurodegeneration. While there are also many vertebrate cell lines
available for analysis, an advantage of many Drosophila cell lines is that they are
highly amenable to RNAi manipulation. Highly efficient target transcript deg-
radation can be achieved with many Drosophila cell lines by simply adding
microgram amounts of dsRNA molecules of 200-400 bp directly to cell culture
media (Boutros
et al.
, 2004; Steinbrink and Boutros, 2008). RNAi could poten-
tially be used to create a Drosophila cell culture model of neurodegeneration, or
to conduct a whole genome screen for dsRNA molecules that modify a cell
culture phenotype. An RNAi screening center, consisting of all of the necessary
resources for conducting genome-wide screens in Drosophila cell lines, was
recently established at Harvard and another recent established in Sheffield
which are currently available for such screens (see http://teratogen.med.har-
vard.edu/ and http://www.rnai.group.shef.ac.uk/). To bring this technology full
circle, the same dsRNAs that make up a “second generation” RNAi library have
been used to generate a publically accessible inducible transgenic RNAi collec-
tion (Vienna Drosophila Resource Centre; Dietzl
et al.
et al.
, 2007). This facility
provides the unprecedented ability to move from
in vitro
screen to
in vivo
analysis
all through community-based initiatives.
B. The neuroanatomy and function of DA neurons in Drosophila
Drosophila has a complex nervous system consisting of approximately 100,000
neurons including a subset of approximately 200 neurons that secrete the neuro-
transmitter DA (Fig. 1.1). The anatomical locations of all of the Drosophila DA
producing neurons have been identified, and their development has been traced
throughout the life cycle. Thus, genetic perturbations affecting the number,
morphology, or locations of DA neurons in Drosophila can be readily identified.
Although the anatomy of the fly brain and the distribution of DA neurons in the
Drosophila central nervous system differ vastly from the vertebrate brain,