Georgiades, Pantelis; (1996) Studies of the expression and function of the retinoid-x-receptor y gene in rodent embryonic development. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Retinoids, such as 9 cis retinoic acid (9cRA) and all trans retinoic acid (tRA), are signalling molecules that are important for the establishment of the correct behaviour of many different types of cells during embryonic development and adult life. Retinoid X receptors (RXRs) and retinoic acid receptors (RARs) are transcription factors that mediate the effects of retinoids by binding to them. RXRs can also be involved in the mediation of non-retinoid signalling molecules because they can form heterodimers with other transcription factors such as thyroid hormone receptors and vitamin D receptors. The objective of the present work was to provide information about the role of RXRγ during rodent development. I isolated a truncated rat RXRγ cDNA clone by screening a rat cDNA library with a chick RXRγ cDNA probe. Using the rat clone as a probe, I examined the expression pattern of RXRγ in rat embryos and found it to be mainly restricted to specific regions of the peripheral and central nervous systems and in skeletal muscle. Its expression in adult rats was also examined and was found to be expressed in specific regions. These include skeletal muscle, heart, brain, sciatic nerve and liver. I also examined the expression of RXRγ in mouse embryos and found it to be essentially the same to that observed in rats. The role of RXRγ in rodent myogenesis was examined by comparing its spatio-temporal expression with that of muscle-specific genes of known function so as to establish the timing of the initiation of RXRγ expression. I showed that the initiation of RXRγ expression was concomitant with muscle differentiation of limb but not early myotomal myoblasts, suggesting that it may be involved in limb myoblast differentiation and that retinoic acid may be a myogenic differentiation signal. This hypothesis was tested in vitro, using a skeletal muscle cell line. As predicted, both 9cRA and tRA were able to relieve the serum-induced inhibition of myoblast differentiation and RXRγ was expressed during this process. RXRγ may mediate the differentiation effect of retinoic acid because its expression was not induced by retinoic acid, since it was expressed in myoblasts in the absence of retinoic acid. The expression of RXRγ in the developing nervous system suggested that it may be involved in the transcriptional regulation of neurone-specific genes. Candidate target genes include those which code for neuronal nicotinic acetylcholine receptor (neuronal nAChR) subunits. The present work supports this because I showed that overexpression of RXRγ in BHK cells activates reporter gene expression from a promoter which is under the control of upstream regulatory sequences derived from the neuronal nAChR ?2 subunit gene, and 9cRA enhances this activation. Moreover, this effect was cell type-specific because when ND7 cells were used, no activation was observed in the presence of RXRγ and 9cRA. This shows that the function of RXRγ can be influenced by trans-acting factors. Finally, I showed that the expression of RXRγ in postnatal heart is age-dependent, suggesting that it may be involved in the regulation of genes whose expression is age-dependent. Moreover, I demonstrated that treatment of primary cultures of neonatal rat cardiomyocytes with either thyroid hormone or tRA leads to changes in RXRγ expression. This finding is consistent with the proposed role of RXRγ because there is evidence implicating these two signalling molecules in cardiac age-dependent changes.

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