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Fig. 2. A cluster of three forkhead-binding sites is sufficient for
dauer-specific lag-2 expression in the IL2 neurons. (A) A
3-kb region upstream of the lag-2 translational start site was
subjected to deletion analysis to determine the minimal fragment necessary for
IL2/dauer-specific lag-2 expression in daf-7 animals. Enzyme
sites used for the generation of the different promoter variants are
indicated: Ns (NspI), A (AccI), E (EcoRI), Nc
(NcoI) and H (HhaI). Solid lines represent fragments of the
lag-2 promoter that were cloned upstream of the GFP-coding sequence;
dashed lines represent deleted sequence. Asterisks represent the location of
the predicted forkhead-binding sites in the lag-2 promoter.
(B) Two potential forkhead-binding sites, named A and B, were
identified in the minimal fragment required for IL2 neuron/dauer-specific
expression. The consensus binding sites for the FoxC1 transcription factor are
indicated in the grey box above the lag-2 sequence (A binding sites).
Capital letters represent the core binding site and small letters indicate
nucleotides required for efficient binding. (C) Smaller deletions of
the 270 bp fragment were created to determine which forkhead-binding sites are
required for IL2 neuron/dauer-specific expression. The white and grey boxes
represent the identified forkhead-binding sites and the crosses indicate
regions where the core binding site sequence was deleted. The relative
intensity of GFP expression in the IL2 neurons is indicated as follows: +++,
strong; ++, moderate; +, faint; -, no expression. For the consensus binding
sites: w can be A or T; m can be A or C; and n can be A, T, C or G.
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