Institute of Biological and Biomedical Sciences

Colin Sharpe

Our current laboratory research

Animal caps injected with noggin and retinoid receptor mRNA

Understanding the role of retinoid signaling in the formation of Xenopus primary neurons

Amphibian and fish embryos produce two nervous systems as they develop. In the frog, Xenopus, a complex nervous system, similar to that found in higher vertebrates, forms over a few months as the tadpole develops and metamorphoses into a froglet.  The embryonic primary nervous system, however, forms rapidly and is active in just a little more than one day following fertilisation. Unlike the complex version, it consists of a small number of large neurones that control simple behaviours associated with the free-swimming lifestyle of the early tadpole. Their small number, large size and rapid differentiation have made the embryonic primary neurones a popular model for vertebrate neurone formation. Embryonic primary neurones are readily identified by the expression of the neural specific type-II beta-tubulin (NST) gene or the peripherin (XIF3) gene. We have found that signaling by retinoic acid is required for the formation of primary neurones.

Retinoic acid (RA) is a derivative of vitamin A that acts as a signaling molecule. It binds the heterodimeric RAR/RXR nuclear receptor proteins, which then activate target gene expression.  When retinoid signaling is inhibited, primary neurones fail to form, in contrast, when elevated, more embryonic primary neurones will form than normal.

A functional analysis of SMRT co-repressor isoforms generated by alternative splicing

A functional analysis of SMRT co-repressor isoforms generated by alternative splicing

In the absence of RA, retinoid receptors interact with the co-repressor SMRT or its paralogue NCoR. This results in the deacetylation of the surrounding chromatin and the active repression of target gene expression. SMRT is a large protein of over 270 kDal and consists of a series of domains and motifs that either coordinate the repressive effects or interact with nuclear receptors. SMRT transcripts are subject to alternative splicing which generates many different isoforms that differ predominantly in the C-terminal domains.  These domains govern interactions with nuclear receptors such as the retinoid receptors.

Analysis of the SMRT gene structure and patterns of SMRT transcript alternative splicing has identified two exons whose pattern of alternative splicing is conserved across a wide range of vertebrates.

Constraining alternative splicing at one of these exons, using antisense oligonucleotides in Xenopus embryos, disrupts development and generates embryos with a disorganised nervous system.  The C-terminal interaction domains of mouse SMRT isoforms have been expressed in collaboration with Prof Kneale’s group.  The isoforms differ in their binding affinities for a range of nuclear receptors.  The combined results suggest that the SMRT isoforms perform different roles in nuclear receptor signaling in the cell.