be achieved either by co-expression of UAS RNAi constructs, or by
reducing gene copy with mutants or deficiencies [21]. To examine
the cellular basis for adult eversion phenotypes one can then use
fluorescent transgenes and immunostaining to highlight key drivers
and markers of the EMT process and examine PE cells in vivo, at the
time of the EMT, or dissect out wing discs and culture them in the
presence of ecdysone, which causes the pEMT to occur ex vivo
(Fig. 1). This method has the advantage that the PE cells are able to
be imaged very clearly, and also provides the option of drug
treatments.
2 Materials
2.1 Drosophila
Stocks
2.1.1 GAL4 Driver Stocks
GAL4 lines that express predominantly in PE cells are listed below
(Table 1). Note that the PE has genetically distinct regions. Aldaz
et al. [15] describe these as a central region of squamous cells,
which lies posterior to the anterior/posterior boundary and a
surrounding region of stripe cells. Tripura et al. [22] call these the
posterior cells and medial-anterior cells, respectively. The PE GAL4
Table 1
GAL4 driver stocks
Stock
Chr Comment
Source
Grunge-GAL4
[24]
III Expresses in the central/posterior region of the PE
[15]
BDSC#6773
puckeredGAL4 [13]
III Although puc-lacZ expresses in the stalk region and
throughout the PE [13], we find that puc-GAL4 is
more restricted to the stalk region
BDSC#6762
UltrabithoraxGAL4 [25]
III Expresses in the central/posterior region of the PE
[25]
L.S. Shashidhara
odd-GAL4
[26]
II Expresses in the medial-anterior/stripe cells [15, 22,
25]
L.S. Shashidhara
pannierGAL4 [27]
III Expresses in the stalk region in both PE cells and DP
cells that will form the central notum
BDSC#3039 and other
stocks
tubP:GAL80
ts
[28]
X,
II,
III
A temperature-sensitive GAL4 repressor, GAL80
ts ,
under the control of the alphaTub84B promoter
BDSC#7017, #7019 and
other stocks
UAS-Dicer2 [23]
X,
II,
III
Co-expression of Dicer-2 can enhance the RNAi
process
BDSC#24648, #24650,
#24651 and other
stocks
118
Sofia Golenkina et al.