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9/8/2010
The role of runt, en, and slp in establishing MP cell fate
Epidermal regulation of runt, en, slp
Wild-type runt4496
odd
en
runt, slp, and en are expressed in distinct patterns in the developing midline
Stage 9
Stage 11
slp
En
Runt
Delta mutant embryos still specify distinct MP fates
slp
En
Runt
Slp1 expression patterns – antibody vs in situ
Slp1 En
Stage 11 Stage 12 Stage 14
Stage 11 Stage 12 Stage 14
Stage 10
slp1 En
MP3 assumes an MP4-like fate in slp1 mutant embryos
Model
Runt Slp En
MP3AMG, MP1 MP4, PMG
tup
Reagent to test this model – Tup – antibody and probe
Tup
tup En
Examine 4 genotypes to test this model
sim-Gal4 uas-slp1
sim-Gal4 uas-enVP16
enA/enE transheterozygote
runt4496 null mutant
Is slp1 sufficient to induce MP3 formation?
Runt Slp En
MP3AMG, MP1 MP4, PMG
SlpSlp
sim-gal4 uas-slp1
tup En
sim-gal4 uas-slp1
ple En
Is tup a downstream target of engrailed?
sim-Gal4 uas-enVP16 enA/enE
Runt Slp En
MP3AMG, MP1 MP4, PMG
tup tup Elav
Is tup a downstream target of engrailed?
tup Odd tup Odd
Runt Slp En
MP3AMG, MP1 MP4, PMG
What is the role of En in posterior neurons?
Does a transcription factor specify MP4 fate?
tup
Hh
What purpose does en serve in midline neurons?
Joe Kearney’s data
Variable number of neurons in en mutant embryos
From 43 segments: Average number of neurons: 7.2 +- 1.9
Range: 4-10
wor Elav
MNB is missing
Average number of Zfh1 positive cells (stage 13-15, 9 segments) : 2.8
Segment mixing?
stage 11
Elav
Zfh1 Elav
tup Elav
Zfh1 Elav
Early enA/enE segments are disorganized
MP1s are also present in en mutant embryos
HbZfh1
tup
MP1smVUMs
H-cell
wor Elav
MNBs
Examine iVUM and mVUM markers – Lim1, vglut, gad1, tkr, and cas
Future en experiments
Rescue En in midline and in paired stripes in an enA/enE mutant background
Ectopically express En and EnVP16 in all midline neurons – consequences?
Drive Runt in midline neurons?
MP1 cell fate
Stage 9
slp1 En
MP1s still form in the absence of either runt or slp1
tup Oddtup Odd
slp12
runt4496
en+ segment en- segment
Look at additional MP1 mutant animals to determine mechanisms that establish MP1 cell fate
hb En
Current Model
Runt Slp En
MP3AMG, MP1 MP4, PMG
tup
odd, hb
Hh