Worm Breeder's Gazette 14(3): 32 (June 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.
Division of Biologcal Sciences, University of Missouri, Columbia, MO 65211
The daf-7 gene encodes a novel member of the TGF-B superfamily. Mutations in daf-7 result in temperature-sensitive (ts) dauer-constitutive (Daf-c) and egg laying (Egl) phenotypes. To further study DAF-7 expression, we have generated integrated lines carrying gfp driven by the daf-7 promoter (daf-7p::gfp) in an N2 genetic background. GFP is strongly expressed in one pair of chemosensory neurons, ASI (we previously reported ASJ, WBG 14.2, 1996), and weakly expressed in one pair of interneurons. GFP expression could be detected about 4 hr after animals hatched in food at 20C. In a daf-7 null genetic background, the transgenic animals (extrachromosomal array) express GFP strongly in ASI and this expression could be detected in dauer larvae formed constitutively at 25C, but was not detected in dauer larvae formed by starvation. These data indicated that daf-7 expression can be regulated by food and pheromone stimuli. Furthermore, functional DAF-7 is not required for maintenance of its own expression in chemosensory neurons. Cell killing experiments (Bargmann and Horvitz, Science 251: 1243, 1991) showed that ASI is one of the chemosensory neurons repressing dauer formation under non-dauer inducing conditions at 20C, but it functions redundantly with ADF and ASG. To test the effect of higher temperature (25C) on dauer formation in cell killing experiments, we ablated ASI alone in the integrated line (daf-7p::gfp). The animals were operated on within 3 hours of being hatched in M9 buffer. Our preliminary results showed that a few operated animals formed dauer larvae in food at 25C. We also transformed N2 into a ts Daf-c mutant by using mec-4(d) expressed under the control of the daf-7 promoter [daf-7p::mec-4(d)] or the srd-1 promoter [srd-1p::mec-4(d)]. The srd-1 gene encodes a candidate chemosensory receptor and is expressed in the ASI neurons of hermaphrodites (Troemel et al., Cell 83: 207, 1995). The transgenic animals also carried daf-7p::gfp to monitor damage of ASI. The animals formed some dauer larvae constitutively at 25!C, and GFP expression was detected in the dauer larvae, indicating ASI was not always killed by MEC-4. However, no more than 70% of animals in the integrated daf- 7p::mec-4(d) line form dauer larvae at 25C, indicating that undamaged neurons such as ADF and ASG function redundantly with ASI. The interneurons weakly expressing daf-7 may also be involved in regulation of dauer formation. Dr. Cori Bargmann helped with laser ablations, neuron identification, and provided srd-1::gfp constructs. Dr. Monica Driscoll provided mec-4(d), Dr. Marty Chalfie provided GFP vector and Dr. Pam Hoppe helped with microinjection.