Worm Breeder's Gazette 17(1): 40 (October 1, 2001)

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.

Understanding the function of egl-38 and lin-48 using heat shock promoters hsp16-2 and hsp16-41

Ashwin Uttam, Helen Chamberlin

941 Bio Science Bldg. Columbus OH. 43210

In C.elegans studies of hindgut development indicate the EGL-38 Pax transcription factor directly regulates lin-48, a gene encoding an Ovo related zinc finger transcription factor. To better understand the function of these two genes, we have made clones that include a heat shock promoter (hsp 16-2 and hsp 16-41) and cDNA for each gene. These clones will allow us to test the effect of ectopic expression for each gene.

            Since egl-38 and lin-48 are initially expressed in embryos, we wanted to test the effect of heat shock on embryos. In preliminary experiments, we observed embryos are very sensitive to the commonly used conditions of 33 ºC for 1+ hours. Consequently, we wanted to identify a heat shock (HS) condition that could both induce heat shock expression and minimize lethality. To identify a good heat shock condition, a gfp cDNA was cloned into the hsp16-2 and hsp 16-41 promoter vectors. After the injection of the animals and the production of transgenic lines, the animals were heat shocked under various conditions (Table 1.1). We found the greatest gfp expression with minimum amount of lethality due to the heat shock was observed when the animals were incubated at 35 º C for half an hour. The above heat shock conditions were also not too detrimental for the animals if they were heat shocked everyday for 24 hours through out their developing life.

Table 1.1

These conditions were then used on transgenic animals carrying a clone with hsp16-2::egl-38, and animals with hsp16-41::lin-48. Although we found neither transgene significantly affected embryonic variability, both affected adult male spicule development (Table 1.2), hsp16-2::egl-38 also affects larvae viability. Genetic results indicate that lin-48 plays an inhibitory role in the development of the male spicules because lin-48 mutant males have ectopic spicule-like structures. The similarity in effect for egl-38 and lin-48 is most likely due to the fact that egl-38 is a transcription factor for lin-48. Therefore, over expressing egl-38 results in an over expression of lin-48, and there by yielding the deformed or absent male spicule phenotype.

Table 1.2