Program of the 2004 East Coast Worm Meeting - Caenorhabditis ...

17. Notch function in differentiated neurons is required to maintain dauer.
Jimmy Ouellet, Richard Roy
Department of Biology, McGill University, Montréal, Québec
Appropriate cell fate specification is highly dependent on cell-cell communication and the
interplay between different pathways provides the developmental complexity typical of higher
animals. In the case of dauer formation in C. elegans, three conserved signaling pathways
(TGF-β, Insulin-like, and cGMP signaling pathways) regulate the complex morphological and
metabolic changes typical of this stage, and many of these changes must occur globally in the
animal. Evidence suggests that these signaling pathways are required in a small subset of
neurons, implicating that a second signal (probably neuroendocrine) is produced in these neurons
to cause these global changes. Although the Notch signaling pathway has not been involved in
dauer formation, we have shown that when the Delta-like ligand (LAG-2) or the Notch receptors
themselves (LIN-12 and GLP-1) are mutated, dauer larvae are unable to maintain this stage and
therefore recover prematurely. When we tested the downstream effector of Notch signaling, the
transcription factor lag-1, for recovery in the daf-7 mutant background we noticed that the animals
maintained the dauer stage better than the single mutant daf-7. This suggests that the
downstream genes activated by Notch, which are required for dauer maintenance, are
’’derepressed’’ in the lag-1mutant background and are rendered independent of Notch signaling.
We also found, through the analysis of the different double mutants, that the Insulin-like pathway
was absolutely required for the premature dauer recovery caused by mutation in components of
the Notch signaling pathway. This suggests that downstream Notch targets may repress
Insulin-like signaling in order to maintain the dauer stage. More interestingly, all these interactions
occur in a small collection of neurons in the head of the animal. We are currently trying to
determine which neurons are involved in Notch signaling during dauer.
Animals harboring a lag-2::GFP transgene express GFP in the IL1 head neurons at the onset
and throughout the dauer stage. Although these neurons were never shown to be required for
dauer development, they are nonetheless among the subset of neurons remodeled during the
dauer stage. To determine if these neurons play a role in dauer development, we used the mutant
background deg-1(u38), which causes the degeneration of the IL1 neurons as well as the
motorneurons AVD, AVG and PVC. We found that these dauer animals are incapable of
maintaining dauer, as was observed in daf-7; lag-2 mutants, suggesting that these neurons are
required for dauer maintenance. Our analysis of the lag-2 promoter indicated that 3 forkhead
binding sites are sufficient for the dauer-specific expression in these head neurons. We are
currently testing the 15 predicted forkhead transcription factors (Fkh) identified from the C.
elegans genome database to determine which Fkh binds to these sites to initiate the Notch
cascade at the onset of dauer. Based on our genetic studies, it appears that the Fkh DAF-16 is
not involved.
We propose that the pathways required for dauer formation (TGF-β and/or the Insulin-like
pathway) activate the Notch ligand in head neurons specifically during dauer through a Fkh.
Ligand binding to the nearby Notch receptors leads to the expression of genes required for the
inhibition of the Insulin-like pathway, probably through a neuroendocrine mechanism since these
changes are rapid and must occur in all cells throughout the animal. Therefore, our results
provide a link between three important signaling pathways, as well as implicating Notch signaling
in a novel function, perhaps different than its more canonical role in binary cell fate specification.