9/8/2010 The role of runt, en, and slp in establishing MP cell fate.

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