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Abstract:
Although biologists have been aware of nematode associations with insects for some time, the details of these interactions have received little attention. A systematic effort to identify the natural ecology of Pristionchus nematodes revealed species-specific host preferences for several beetle species, including the oriental beetle found in Japan and northeastern U.S. In Pristionchus pacificus, the first and only molecular component identified so far that is involved in odor signaling is the cGMP dependent protein kinase, Ppa-EGL-4. To obtain more upstream factors involved in insect pheromone attraction, we performed a non-saturating X-ray mutagenesis on P. pacificus for mutants that do not show attraction toward their host oriental beetle pheromone (ZTDO) using the conventional plate chemotaxis assay. Of the seven obi candidates isolated, we selected two lines for further analyses due to their strong obi chemotaxis phenotype, viability, as well as linked morphological phenotypes after >3x outcrossing to wildtype. The two promising candidates were named obi-1 and obi-3 for Oriental Beetle pheromone Insensitive mutants. The chemosensory phenotype of obi-1 and obi-3 is specific for ZTDO and does not affect the chemoattraction toward other known P. pacificus attractants such as the plant volatile β-caryophyllene and the moth pheromone ETDA. Neither obi-1 nor obi-3 show dauer formation defect (daf-c) or temperature sensitivity. However, both obi-1 and obi-3 animals show higher frequency and amplitude turns than wildtype on OP50. Adult obi-1 hermaphrodites are ~20%; longer than wildtype, with a body length-to-width ratio of 17 compared to 13 in the wildtype. An increase in egg retention (egl) also accompany this long phenotype (lon). Because the chemotaxis assay with cGMP treatment is extremely labor intensive, we used a linked long body morphological phenotype in obi-1 to conduct subsequent mapping of F2 progeny from crosses between obi-1 and the mapping strain Washington. Using 200 F2 lines and molecular markers (Single Stranded Conformational Polymorphisms and Simple Sequence Length Polymorphisms), we were able to confine obi-1 to a 153 kb region represented by 10 subcontigs of Supercontig 58. No known C. elegans long mutants map near the syntenic chromosome region in P. pacificus. Obi-1’s odor specific phenotype and genetic location suggest that it is not an allele of egl-4 (Chr IV) and thus likely to be a parallel or upstream component of odor signaling in P. pacificus.
