Cloning and expression of arylalkylamine N-acetyltranferase-2 during early development and metamorphosis in the sole Solea senegalensis

Oct 29, 2008General and comparative endocrinology

Production and activity of a key melatonin-making enzyme during early growth and transformation in the sole fish

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Abstract

A significant 60-fold reduction in Aanat2 expression was observed just before metamorphosis in Solea senegalensis.

AANAT2 is a key enzyme in the production of melatonin, which varies in expression during development.
Sole larvae showed specific AANAT2 expression in the pineal gland starting from 2 days post-fertilization.
The expression of Aanat2 was higher during the dark phase compared to the light phase at most developmental stages.
The highest levels of AANAT2 mRNA were recorded at 2 and 4 days post-fertilization.
A novel inverse relationship between AANAT expression and thyroid hormone levels was identified during metamorphosis.

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The arylalkylamine N-acetyltransferase (AANAT) is a key enzyme in the rhythmic production of melatonin. Two Aanats are expressed in teleost fish, one retinal specific, Aanat1, and the other one pineal specific, Aanat2, being the latter the main enzyme responsible of the plasma nocturnal melatonin increase in fish. In anurans melatonin has been involved in metamorphosis through antagonizing thyroid hormone function; however, no available data reports a relationship between melatonin system and metamorphosis in fish. In this study, we have cloned the AANAT2 (SsAanat2) in a flatfish, Solea senegalensis, and studied its sites of expression and developmental expression pattern by in situ hybridization and Real Time PCR. These studies allowed us to demonstrate a specific signal in the pineal gland of sole larvae from 2 days post-fertilization (dpf), which was evident until post-metamorphosis. Immunohistochemical analysis on the hybridized slides showed that the sole pineal Aanat2 expressing cells corresponded to pineal photoreceptor cells. Real Time PCR was performed in animals kept under natural photoperiod and sampled at different stages from 0 to 21 dpf (including pre-, early-, middle- and late-metamorphic stages) and at midlight (ML) and middark (MD) daytimes. Sole Aanat2 expression was higher at MD than at ML from 2 dpf and at most developmental stages analyzed. The highest AANAT2 mRNA abundance was observed at 2 and 4 dpf. A significant 60-fold reduction in Aanat2 expression was seen just before metamorphosis demonstrating, for the first time in a vertebrate species, that the expression of pineal AANAT and thyroid hormones levels exhibit an inverse pattern during metamorphosis.

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Cloning and expression of arylalkylamine N-acetyltranferase-2 during early development and metamorphosis in the sole Solea senegalensis

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