Qualifications:
B.Sc. Applied Biochemistry (Yr in Industry): First Class Honours (2003)
School of Biological Sciences, University of Liverpool.
Research
Project:
The Biological Role of Natural Antisense Transcripts
Natural antisense transcripts (NATs) are transcripts encoded from the so-called non-sense DNA strand and exhibit complementarity to transcripts encoded from the sense strand i.e. bi-directional transcription produces overlapping but oppositely orientated transcripts capable of forming hybridization pairs. NATs can be divided into classes based upon the loci from which they originate.
cis-encoded NATs (regarded as “true/real” natural antisense transcripts) are encoded from the same gene locus as their sense counterparts, and show high levels of complementarity (formation of perfect hybrid duplexes); whereas, trans-encoded NATs originate from distinct loci separate from their sense partners, and show a lower level of complementarity (less than 100bp of overlap giving rise to imperfect hybridization duplexes).
There has been considerable speculation regarding the function, if any, of natural antisense transcripts at a molecular and physiological level, with some regarding antisense transcription as nothing more than background transcriptional “noise”, and others assigning a major role in gene regulation to these transcripts.
The effect of antisense transcripts on gene expression and cellular phenotype can be illustrated best by the results of experiments using synthetic oligonuleotides or antisense RNA expression vectors. Whereby, Gene silencing can be induced by synthesizing an oligonuleotide that is complementary (i.e. antisense) to a region of the gene of interest; or by the expression of an antisense RNA from an engineered plasmid vector.
It would be particularly reckless for scientists to think that these “man-made” tools were not already employed by nature with even more intricate and elegant methodologies, utilizing natural antisense transcripts for gene regulatory purposes. There have been both prokaryotic and eukaryotic examples of natural antisense transcripts identified as playing key roles in gene regulation.
In the Werner lab we are using the Zebrafish (Danio rerio) model system to elucidate the mechanism(s) utilised by the cell for natural antisense–mediated gene regulation. Specifically, we are using the sodium phosphate co-transporter gene locus as our model system in the Zebrafish.
An antisense transcript related to the sodium dependent inorganic phosphate cotransporter (NaPi-II) was first identified in 1992 by Werner et al in winter flounder (Pleurontectes americanus) as an unexpected northern blot product. At the time of its discovery the 624 bp antisense transcript was thought to encode a peptide of 68 amino acids, this was later disproved.
The identification of a related antisense transcript in Zebrafish (Danio rerio) showed an antisense transcript comprising five sense-related exons, with the fifth one being alternatively spliced. The longer antisense transcript spans the whole of the sense protein coding gene. A schematic representation of the Zebrafish npt2b1 gene locus is shown below:
The major aims of the project are:
• To characterise the expression profiles of the sense and antisense transcripts
of the npt2b1 gene locus.
• To elucidate the mechanism by which the antisense transcript controls
the sense transcript expression.
• Apply the mechanistic model to other antisense controlled systems.
An overview of the project is given below:
