C. elegans secrete a synergistic blend of derivatives of the dideoxysugar ascarylose, which promote entry into dauer diapause, an alternate non-feeding larval stage that is highly stress-tolerant and can persist for many months (Butcher et al., 2007). It has been speculated that this dauer-inducing signal (the dauer pheromone) is secreted in response to unfavorable environmental conditions; however, concrete evidence that any of the ascarosides are produced in response to stress has been lacking. Here we show that of the two major components of the dauer pheromone, ascr#2 and ascr#3, ascr#2 is strongly induced by starvation, whereas production of ascr#3 remains largely unaffected. Content of ascr#2 and ascr#3 in supernatant and worm pellet from starved and non-starved liquid cultures was assessed using Differential Analyses by 2D-NMR Spectroscopy (DANS) (Schroeder et al., 2007). These NMR-spectroscopic analyses showed that non-starved wildtype (N2) and
hlh-13(
tm2279) cultures produced large quantities of ascr#3, but did not produce detectable quantities of ascr#2. On the other hand, starved cultures consistently produced large amounts of both ascr#2 and ascr#3. For wildtype worms (N2) and
hlh-13(
tm2279) mutants, it appears that production of ascr#2 is at least 100-fold upregulated in response to starvation. In contrast,
daf-9(
m540) and
daf-9(
dh6):
daf-12(
rh411,
rh61) did not produce ascr#2 under either starved or non-starved conditions. Ascr#3 production appeared unchanged in these two mutant strains. Our results fit well with the observation that ascr#2 is the more potent dauer-inducer, whereas ascr#3 is more active as a sex pheromone (Srinivasan et al., 2008). Additional studies suggest that ascr#2 also affects thermotolerance of adult worms specifically under starvation conditions.