Investigation of Splicing Factors Regulating IR Isoform DAF-2B Generation in C. elegans
In the nematode C. elegans, an alternatively spliced, non-signaling insulin receptor (IR) isoform DAF-2B sequesters insulin peptides to modulate insulin sensitivity, altering animal physiology and development. Associated with reduced insulin signaling is entry to the dauer diapause, an alternative survival state enabling longevity under adverse environmental conditions. Genetic screens have previously identified splicing factors that increase or decrease daf-2b expression upon knockdown; however, understanding of splicing factor regulation of phenotypic effects through dauer entry were to be investigated. To approach this question, RNA interference was used to deplete gene products of candidate splicing factors though feeding C. elegans bacteria expressing complementary double stranded RNA. Exposure to increased temperature provided the adverse environmental stimulus for dauer entry. Total number of dauers and non-dauers were recorded through a qualitative assay. Upon measurement of dauer entry rate related to knockdown of selected candidate splicing factors, 8 out of the 15 clones tested were found to significantly alter dauer formation. Seven clones (rsp-2, prp-31, asd-2, usp-39, prp-21, snu-23 and sfa-1) showed increases in daf-2b expression while only mec-8 corresponded to decrease. Plotting rates against known DAF-2B splicing effects, the results supported a general positive correlation between daf-2b expression and dauer formation. Overall, the results of the experiment suggests that splicing factors play a role in DAF-2B expression resulting in an altered insulin signaling. Further insight into candidate splicing factor clones causing significant effects on dauer formation may prove beneficial to increasing understanding of DAF-2B’s role in insulin signaling related to aging.