Worm Breeder's Gazette 14(2): 36 (February 1, 1996)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Expression of DAF-7 in chemosensory neurons

Peifeng Ren(ren@dauerdigs.biosci.missouri.edu), Patrice S. Albert , Donald L. Riddle

Division of Biological Sciences, University of Missouri-Columbia, MO 65211 Mutations in daf-7 result in constitutive formation of dauer larvae. The daf-7 gene encodes a secreted protein that is a novel member of the TGF- beta superfamily (Chang-Su Lim, Ph.D. Thesis, 1994). Laser ablation experiments have determined that amphid sensory neurons ADF, ASG, ASI and ASJ function in the regulation of dauer formation (Bargmann et al., Science 251,1243). Our hypothesis is that daf-7 might be expressed in the amphid neurons to promote non-dauer development in response to food stimuli. Secreted DAF-7 ligand may then bind the DAF-1 and/or DAF-4 receptors which are transmembrane receptor serine/threonine kinases in the TGF-beta receptor family (Georgi et al., Cell 61, 635; Estevez et al.,

Nature 365, 644). Our expression data indicated that daf-1 and daf-4  are 
expressed in interneurons and in amphid neurons (Georgi and Estevez, 

To study the expression pattern of daf-7, we have generated transgenic 
worms carrying a gfp reporter gene expressed under the control of the 3 
kb daf-7 promoter region from the rescuing genomic DNA fragment (daf-
7p::gfp). In an N2 genetic background, we found that GFP was expressed in 
one pair of amphid neurons, ASJ. GFP expression can be detected in the 
four larval stages and adults, but not in eggs or in dauer larvae formed
in pheromone or by starvation. During dauer recovery, GFP expression was 
detected after the dauer larvae were placed in food. Dauer-inducing 
pheromone suppressed the GFP expression. These data suggested that daf-
7 expression can be regulated by food and pheromone stimuli.

The dominant allele of mec-4 , which causes swelling and death of the 
touch receptor neurons, has been used to define the function of other 
neurons (Maricq et al., Nature, 378,78). We placed mec-4(d), encoding  
degenerin, under the control of the daf-7 promoter [daf-7p::mec-4(d)].
This construct, coinjected with rol-6, transformed N2 into a temperature-
sensitive(ts) dauer-constitutive (Daf-c) mutant. In synchronous 
populations,  1%, 5% and 28% of the rollers formed dauers at 15 C, 20 C 
and 25 C, respectively.  However, these dauer larvae recovered 
spontaneously at 25 C. Since the laser ablation results indicated that ASJ
is required for efficient dauer recovery, we again transformed N2 into a
ts Daf-c mutant, this time carrying both daf-7p::gfp and daf-7p::mec-4(d).

We found GFP expression in dauer larvae, indicating there is sufficient 
function to allow dauer larvae to recover. The MEC-4 activity may result 
in sufficient loss of DAF-7 secretion from ASJ to cause entry into the 
dauer state at 25 C; but in the absence of high pheromone the gfp gene 
was still expressed. Since in the transgenic animals the expression of 
mec-4(d) and native daf-7 are driven by daf-7p  simultaneously, some 
animals could have made enough DAF-7 to promote development to the 
adult before ASJ was sufficiently damaged. This, plus some somatic loss 
of the transgene, may account for the low percentage (28%) of dauer 
formation by transgenic animals at 25 C. The daf-7 mutants are all ts, 
but they form 100% dauers at 25 C. We conclude that ASJ is involved both 
in preventing entry into the dauer stage at higher growth temperatures 
and in exiting from the dauer stage in response to food stimuli.

Dr. John White helped with identifying neurons, Dr. Monica Driscoll 
provided mec-4(d) ,  Dr. Marty Chalfie provided GFP vector and Dr. Pam 
Hoppe helped with microinjection.