The tailless family of nuclear receptors is highly conserved among animals. In humans, it functions in regulating neuronal stem cell differentiation. The C. elegans tailless ortholog,
nhr-67, is expressed in a dynamic pattern in pre-uterine cells: initially in the 4 pre-VU cells during the L2, then upregulated in the anchor cell (AC) in response to the
lin-12/lag-2 Notch reciprocal signaling system. During the L3 stage,
nhr-67 expression is maintained at high levels in the AC and at low levels in VU descendants that produce the adult ventral uterus.
nhr-67 is required for expression of the
lin-12/Notch receptor in pre-VU and VU cells and for multiple markers of AC identity, indicating that it functions in differentiation of both uterine cell types. Loss of
nhr-67 in the AC and pre-VU cells leads to a loss of
lag-2 expression in the AC and a failure of the AC to exit the cell cycle, indicating that differentiation of the AC is compromised. Deletion of a 276bp region of the
nhr-67 promoter results in a loss of
nhr-67 expression in pre-VU, AC, and VU cells. A 160bp enhancer that includes eight conserved candidate cis elements is necessary and sufficient for
nhr-67 expression during ventral uterine development. The region includes two E box sequences that we propose bind the HLH-2 transcription factor, which functions in AC and pre-VU development. Deletion analysis and site-directed mutagenesis experiments indicate that homodimeric HLH-2 binding sites are required for AC and pre-VU expression, but loss of
hlh-2 compromises reporter gene expression in the AC, but not pre-VU cells. Our data demonstrate the primary role of the E box sequences in regulating
nhr-67 in the AC and pre-VU cells; the former apparently via HLH-2 homodimers and the latter via either another HLH protein or HLH-2 heterodimer. Supported by the NIGMS.