Enhanced at puberty 1 (EAP1) is a new transcriptional regulator of the female neuroendocrine reproductive axis
Sabine Heger, … , Wolfgang Sippell, Sergio R. Ojeda
Sabine Heger, … , Wolfgang Sippell, Sergio R. Ojeda
Published August 1, 2007
Citation Information: J Clin Invest. 2007;117(8):2145-2154. https://doi.org/10.1172/JCI31752.
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Enhanced at puberty 1 (EAP1) is a new transcriptional regulator of the female neuroendocrine reproductive axis

Abstract

The initiation of mammalian puberty and the maintenance of female reproductive cycles are events controlled by hypothalamic neurons that secrete the decapeptide gonadotropin-releasing hormone (GnRH). GnRH secretion is, in turn, controlled by changes in neuronal and glial inputs to GnRH-producing neurons. The hierarchical control of the process is unknown, but it requires coordinated regulation of these cell-cell interactions. Here we report the functional characterization of a gene (termed enhanced at puberty 1 [EAP1]) that appears to act as an upstream transcriptional regulator of neuronal networks controlling female reproductive function. EAP1 expression increased selectively at puberty in both the nonhuman primate and rodent hypothalamus. EAP1 encoded a nuclear protein expressed in neurons involved in the inhibitory and facilitatory control of reproduction. EAP1 transactivated genes required for reproductive function, such as GNRH1, and repressed inhibitory genes, such as preproenkephalin. It contained a RING finger domain of the C3HC4 subclass required for this dual transcriptional activity. Inhibition of EAP1 expression, targeted to the rodent hypothalamus via lentivirus-mediated delivery of EAP1 siRNAs, delayed puberty, disrupted estrous cyclicity, and resulted in ovarian abnormalities. These results suggest that EAP1 is a transcriptional regulator that, acting within the neuroendocrine brain, contributes to controlling female reproductive function.

Authors

Sabine Heger, Claudio Mastronardi, Gregory A. Dissen, Alejandro Lomniczi, Ricardo Cabrera, Christian L. Roth, Heike Jung, Francesco Galimi, Wolfgang Sippell, Sergio R. Ojeda

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Figure 3

EAP1 is a nuclear protein with dual transcriptional activity expressed in hypothalamic neurons that facilitate or inhibit the initiation of female puberty.

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EAP1 is a nuclear protein with dual transcriptional activity expressed i...
(A) Confocal image of a subregion of the rat POA from a 28-day-old female rat showing that EAP1 has a predominant nuclear localization (green). Notice that not all neurons contain EAP1, as evidenced by the lack of green staining in cell nuclei identified by Hoechst staining (blue). Long arrows indicate EAP1-positive cells; short arrows indicate cells lacking EAP1. (B) Detection of EAP1 (green) in the nucleus of GnRH neurons (red, arrows). (C) Wide-field image of a neuron labeled for EAP1 and Hoechst after constrained iterative deconvolution and 3D reconstruction showing that EAP1 (red) is not associated with condensed chromatin (blue). Scale bars: 40 μm (A); 20 μm (B); 5 μm (C). (D–F) Combined immunohistochemistry–in situ hybridization of brain sections from 28- to 30-day-old female rats showing that EAP1 protein is abundant (D) in cells of the latero-ventral portion of the ventromedial nucleus (LVMH) of the hypothalamus, identified as enkephalinergic by their content of preproenkephalin (preproEnk) mRNA (E and F). The higher-magnification image in F shows that all preproenkephalin mRNA–containing neurons (white grains) are EAP1 positive (brown staining; examples denoted by arrows). 3V, third ventricle. (G–I) EAP1 has a dual transcriptional regulatory activity that requires an intact RING finger domain. (G) In GT1-7 GnRH-producing cells, EAP1 transactivates the GnRH promoter. (H) In the same cells, it represses the preproenkephalin promoter. (I) This repressive activity is also seen in hippocampal neuroprogenitor HiB5 cells. Both the transactivating and repressive activities of EAP1 are abolished by either a single amino acid substitution (C→A) in the EAP1 RING finger domain or by deletion of this domain (ΔR). Numbers above bars represent number of wells/group, and error bars are SEM.