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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Research Article Endocrinology

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 5

In vivo lentivirus-mediated delivery of EAP1 shRNAs to the rat POA decreases EAP1 protein levels.

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In vivo lentivirus-mediated delivery of EAP1 shRNAs to the rat POA decre...
The images illustrate results derived from 8 control animals injected with LV eGFP, 6 animals correctly injected with EAP1 sh1, and 2 animals with misplaced injections of EAP1 sh1. (A and B) Examples of well-placed injections carrying EAP1 sh1–encoding lentiviruses. The cells infected are seen along the lateral edge (A) or dorsal-medial aspect (B) of the AVPV. (C) Example of misplaced injections (too lateral and too caudal). (D) Higher-magnification image showing that most infected cells had neuronal morphology. (E) Example of an infected GnRH neuron (yellow). (F–I) EAP1 protein content (red) was reduced in neurons infected with EAP1 sh1 (green). (F) The injection track, rich in eGFP-expressing neurons, shows fewer EAP1-positive cells (red) than neighboring areas. (G) Higher-magnification images illustrating the reduction in EAP1 immunoreactive material (red) in the nucleus of EAP1 sh1–infected cells (eGFP-positive, green) as compared with noninfected (eGFP-negative, red only) cells. (H) Same field as in G, but showing only EAP1 immunoreactivity (red) to illustrate the loss of EAP1 protein in EAP1 sh1–infected cells. (I) Fluorescence intensities in AUs based on measurements of 36 infected and 104 noninfected cells. ***P < 0.01 versus noninfected cells. Scale bars: 20 μm (D); 5 μm (E); 50 μm (F); 10 μm (G and H). Numbers in parentheses represent number of wells per group, and error bars are SEM. Arrows in A–C point to sites of injection; arrows in G and H point to infected cells (G) exhibiting a decrease in EAP1 immunoreactivety (H).