Homeobox genes and melatonin synthesis: regulatory roles of the cone-rod homeobox transcription factor in the rodent pineal gland

Abstract

Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT) enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a cAMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processes, are also expressed in the mature rodent pineal gland. Among these, the cone-rod homeobox (CRX) transcription factor is believed to control pineal-specific Aanat expression. Based on recent advances in our understanding of Crx in the rodent pineal gland, we here suggest that homeobox genes play a role in adult pineal physiology both by ensuring pineal-specific Aanat expression and by facilitating cAMP response element-based circadian melatonin production.

Figure 1

Appearance of Otx2, Crx, and Aanat transcripts in the developing rat pineal gland. Otx2 and Crx expression peak around birth [14], at which time Aanat expression is initiated [19]. The spatial and temporal correlation between peaks in expression of Otx2 and Crx and the start of Aanat expression supports that the OTX2 and CRX homeodomain transcription factors induce Aanat transcription [7]. E, embryonic day; P, postnatal day.

Figure 2

Crx and Aanat day/night expression in the adult rat pineal gland. Radiochemical in situ hybridization autoradiographs of midsagittal brain sections through the pineal gland of rats killed at daytime (Zeitgeber time 6) and nighttime (Zeitgeber time 18). Images are orientated with caudal to the left. Crx displays a specific expression in the pineal gland. Studies have shown a diurnal rhythm of Crx expression in the rat pineal gland with a night time peak [7, 35, 71]. Aanat also exhibits a pineal-specific expression; however, daytime transcript levels are very low. Scale bar, 1 mm. Methodological details have been previously published [14, 71].

Figure 3

Working model of the role of homeobox genes in generation of pineal specificity and circadian output. CRX and OTX2 homeodomain transcription factors and NE/cAMP/CREB signaling, initiated by NE released from the sympathetic nerve endings at night, act on PCE and CRE cis-regulatory elements in the pineal Crx promoter region, respectively, to generate tissue-specific circadian expression of Crx. CRX protein acts on PCEs in the Aanat promoter region and confers a pineal-specific and circadian expression of Aanat. The homeodomain transcription factor generated rhythmicity of Aanat transcription supports the classical NE/cAMP/CREB-driven circadian rhythm of Aanat expression. The Crx promoter contains at least one PCE and several CRE cis-regulatory elements. In the Aanat promoter, several CREs and at least two PCEs exist. Promoter analysis was performed using Genomatix MatInspector software [72]. CRE, cAMP response element; CREB, cAMP response element binding protein; PCE, photoreceptor conserved element.