The impact of Alu elements on the human proteome

Approximately 45% of the human genome is comprised of mobile, or transposable,
DNA elements (TEs). Of this, 11% is attributed to Alu elements. Alu elements are
approximately 300 base pairs in length and are primarily located in the introns of
non-coding DNA. However, in some cases, the introduction of an alternative splice
site, as a result of an Alu insertion in a protein-coding region, leads to the exonisation
of a partial Alu sequence. This exonisation can lead to the expression of an
alternative protein isoform which may have disrupted or altered function and
therefore, could have the potential to be cause disease.
Through the use of bioinformatics, this project firstly aimed to predict the extent of
Alu exonisation and subsequent translation in the human proteome. Additionally,
through the use of local sequence alignments, the nature of observed insertions could
also be studied. Once prior aims were established, a series of techniques were used
to study the possible effects of translated Alu insertions on the structure and function
of proteins. A number of protein variants were expressed and purified from E. coli.
Using biophysical techniques, such as ITC and CD, Alu structure and any effects of
Alu insertions on the ligand binding and stability of MBP were studied. Additional
binding experiments were performed as a means to explore a potential binding
interaction between an Alu-like sequence with geldanamycin, an interaction which
was initially observed using phage display.
A secondary avenue of research was performed in collaboration with the Aspden
and Wurdak groups at the University of Leeds to investigate the difference in
translation levels of ‘Alu’ and ‘non-Alu’ mRNAs in human cells. Analysis was
performed using a combination of polysome profiling, reverse transcription and
quantitative PCR.