Biochemistry

Motifs in mRNAs that control protein synthesis:

The identification of hidden signals in mRNA sequences

These projects involve the use both computer (bioinformatic) and experimental tools to test for new translation mechanisms. Bioinformatics was recently highlighted as a key field of expertise for the life sciences in the future. It arose at the interface of the fields of biology, mathematics and computer science to permit interpretation of large quantities of biological data, particularly DNA sequences. The recent availability of large amounts of sequence data, particularly complete sequences, has revolutionised the study of many organisms.  This work is part of the Otago University Area of Research Excellence entitled 'Bioinformatics and Computational Biology'  

This study takes the approach of inferring the biology of translation from the DNA blueprint, then testing the ideas experimentally in tissue culture cells. New assays have been developed which utilise the power of modern microscopic and cell biological techniques. The elements discovered  include stability/instabilty elements (e.g. AU rich elements) and those for mRNA localisation in mammalian cells.

This research is facilitated by a database of translational control elements

TransTerm is a database of sequence contexts about the stop and start codons of many species found in GenBank. TransTerm also contains codon usage data for these same species and summary statistics for the sequences analysed. A database of translational control elements has been established an included in our TransTerm database. Other Software and Web servers, Datasets from the Brown Lab.

Translational Control in of Hepatitis B protein synthesis

Viruses frequently use unusual mechanisms to make essential proteins. Commonly these involve translational control or 'recoding' mechanisms. These are potential sites for anti-viral agents. Most of these recoding mechanisms are poorly understood, but these events are usually directed by sequences and structures in the viral RNAs. These signals may be short adjacent RNA sequences or more complex structures. The synthesis of HBV polymerase (P) protein involves a complex process that differs from host cell protein synthesis, and is a potential target for antiviral agents. The P protein translation initiation site (AUG) is preceded by multiple potential initiation sites, to make any P protein these must somehow be bypassed. P protein is essential for HBV maintenance and multiplication. A new hypothesis for P protein synthesis will be tested. The study will involve testing the expression of reporter constructs containing part of the viral genome in animal cells. This study will identify the key components of this process, and may suggest new avenues for treatment of HBV targeted at this mechanism.

A new method to identify the sites of RNA-protein interactions in living cells.

Protein expression depends significantly on the stability, translation efficiency and localization of mRNA. These qualities are largely dictated by the RNA-binding proteins associated with an mRNA. We have developed a new method to visualize and localize RNA-protein interactions in living mammalian cells. This methods uses trimolecular fluorescence complementation assay (TriFC) to locate RNA-protein interactions where they occur in living cells using confocal microscopy.

Using this method, we found that the fragile X mental retardation protein (FMRP) isoform 18 and the human zipcode-binding protein 1 ortholog IMP1, an RNA transport factor, were present on common mRNAs. These interactions occurred predominantly in the cytoplasm, in granular structures. In addition, FMRP and IMP1 interacted independently of RNA. Tethering of FMRP to an mRNA caused IMP1 to be recruited to the same mRNA and resulted in granule formation. The intimate association of FMRP and IMP1 suggests a link between mRNA transport and translational repression in mammalian cells (Rackham and Brown, 2004).

Selected Publications

SJ Lange, D Maticzka, M Mohl, JN Gagnon, CM Brown and R Backofen (2012) Global or local? Predicting secondary structure and accessibility in mRNAs. Nucleic Acids Research (Feb 2012 in press) (web resources)

Chen, A., and Brown, C. (2012) Distinct families of cis-acting RNA replication elements epsilon from hepatitis B viruses. RNA Biology 2012: 2 (published online 25/12/2011)

Stevens, S.G., Gardner, P.P., and Brown, C. (2011) Two covariance models for iron-responsive elements. RNA Biology 8: 792-801. (web resources)

Panjaworayan, N, S Payungporn, Y Poovorawan, CM Brown (2010) Identification of an effective siRNA target site and functional regulatory elements, within the hepatitis B virus post-transcriptional regulatory element. Virology J 7:216

Milev, M, CM Brown and AJ Mouland (2010) Live cell visualization of the interactions between HIV-1 Gag and the cellular RNA-binding protein Staufen1 Retrovirology 7:41.

Jacobs, G.H., A. Chen, S.G. Stevens, P.A. Stockwell, M.A. Black, W.P. Tate, and CM Brown (2009) Transterm: a database to aid the analysis of regulatory sequences in mRNAs. Nucleic Acids Res, 37:D72-76. (web resources)

Panjaworayan, N, SK Roessner, AE Firth and CM Brown (2007) HBVRegDB: Annotation, comparison, detection and visualization of regulatory elements in hepatitis B virus sequences, Virology J 4:136 (web resources)

Zadissa, A, JC McEwan and CM Brown (2007) Inference of transcriptional regulation using gene expression data from the bovine and human genomes, BMC Genomics 2007, 8:265

Firth, A and Brown CM. (2006) Detecting overlapping coding sequences in virus genomes BMC Bioinformatics 7:173

GH Jacobs, PA Stockwell, WP Tate and Brown, CM (2006) Transterm—extended search facilities and improved integration with other databases Nucleic Acids Res. 34:D37-D40

Chung B.Y.W., C. Simons, A. E. Firth, C.M. Brown and R. P. Hellens (2006) Effect of 5'UTR introns on gene expression in Arabidopsis thaliana. BMC Genomics 7:120

Chen, A., Y.F. Kao, and C.M. Brown (2005) Translation of the first upstream ORF in the hepatitis B virus pregenomic RNA modulates translation at the core and polymerase initiation codons. Nucleic Acids Res, 2005. 33:1169-1181.

Firth, A and Brown, CM (2005) Detecting overlapping coding sequences with pairwise alignments. Bioinformatics 21:282-292

Chapman, B and Brown, CM (2004) Translation termination in A. thaliana: characterisation of three versions of release factor 1. Gene 341:219-225

Rackham, O and Brown, CM (2004) Visualization of RNA-protein interactions in living cells: FMRP and IMP1 interact on mRNAs EMBO J 23:3346-3355

Brown, C. M. Jacobs, G, Stockwell, S and Schreiber, M. (2003) Detection of Signals in mRNAs that influence translation Applied Bioinformatics 2:S47-52

Schreiber, M. C. M. Brown (2002) Compensation for nucleotide bias in a genome by representation as a discrete channel with noise. Bioinformatics 18:507-512

Jacobs, G. H., O. Rackham, P. A. Stockwell, W. Tate C. M. Brown (2002) Transterm: a database of mRNAs and translational control elements. Nucleic Acids Res 30: 310-1.

Jacobs, G.H., Stockwell, P., Schreiber, M., Tate, W.P. and Brown, C.M. (2000) Transterm: a database of messenger RNA components and signals. Nucleic Acids Res 28:293-295

Brown, C.M., G.Jacobs, M.Schreiber, J.Magnum, J.McNaughton, M.Cambray, M.Futschik, L.Major, O.Rackham, W.Tate, P.Stockwell, C.Thompson, N.Kasabov (1999) Using bioinformatics to investigate post-transcriptional control of gene expression, NZ BioScience 7:11-12

Dalphin, M.E., P.A. Stockwell, W.P. Tate, & C.M. Brown (1999) TransTerm, the translational signal database, extended to include full coding sequences and untranslated regions. Nucleic Acids Res. 27:293-294.

Dalphin, M.E., C.M. Brown, P.A. Stockwell, W.P. Tate (1998) Transterm is now a relational database. Nucleic Acids Res. 26:335-337.

Miller, W.A., Brown, C.M. and Wang, S.P. (1997) New punctuation in the genetic code: luteovirus gene expression. Seminars in Virology 8:3-13

Dalphin, M.E., Brown, C.M., P.A. Stockwell and Tate, W. P. (1997) The translational signal database, TransTerm: more organisms, complete genomes. Nucleic Acids Res. 25:246-247

Brown, C.M., Dinesh-Kumar, S.P. and Miller, W.A. (1996) Local and Distant Sequences are Required for Efficient Read-Through of the Barley Yellow Dwarf Virus-PAV Coat Protein Gene Stop Codon J. Virol. 90:5244-5250

Dalphin, M.E., Brown, C.M. Stockwell, P.A. and Tate, W.A. (1996) Transterm: The Translational Termination Database Nucleic Acids Res. 24:216-218

Brown, C.M., Quigley, F.R., Miller, W.A. (1995) Three eukaryotic release factor one (eRF1) homologs from Arabidopsis thaliana Columbia (U40217, U40218, X69374, X69375). Plant Physiol. 110:335

McCaughan, K.K. †, Brown, C.M.†, Dalphin, M.E., Berry, M. J. and Tate, W. P. (1995) Translational termination efficiency in mammals is regulated by the base following the stop codon. Proc. Natl. Acad.Sci USA 92:5431-5435 † Joint first authorship

Poole, E. S., Brown, C.M. and Tate, W.P. (1995) The Identity of the Base Following the Stop Codon Determines the Efficiency of in vivo Translational Termination in Escherichia coli. EMBO J. 14:151-158

Brown, C.M. and Tate, W.P. (1994) Direct Recognition of mRNA Stop Signals by Escherichia coli Polypeptide Chain Release Factor Two. J. Biol.Chem. 21:33164- 33170

Brown, C.M. , Stockwell, P. A., Dalphin, M. E. & Tate, W. P. (1994) The Translational Termination Signal Database (Transterm) now Includes Initiation Contexts Nucleic Acids Res. 22: 3620-3624

Brown, C.M., McCaughan, K.K. & Tate, W.P. (1993) Two Regions of The Escherichia Coli 16S rRNA Are Important For Decoding Stop Signals In Polypeptide Chain Termination. Nucleic Acids Res. 21: 2109-2115.

Tate, W.P. & Brown, C.M. (1992) Translational Termination: Stop for Protein Synthesis or Pause for Regulation of Gene Expression Biochemistry 31: 2443-2450.

Brown, C.M., Stockwell, P.A. Trotman, C.N.A. & Tate, W.P. (1990) Sequence analysis suggests that tetra-nucleotides signal the termination of protein synthesis in eukaryotes. Nucleic Acids Res. 18: 6339-6345.

Brown, C.M., Stockwell, P.A. Trotman, C.N.A. & Tate, W.P. (1990) The signal for the termination of protein synthesis in prokaryotes. Nucleic Acids Res. 18: 2079-2085.

Thanks to current and past funding agencies:

Dunedin School of Medicine Bequest Funds
Otago Research Grant
Human Frontier Science Organisation
Lotteries Health
The Marsden Fund

Other Projects

I am interested in contacts from potential collaborators, post-doctoral fellows, and graduate students. Information on fellowships and scholarships can be found on the University of Otago Postgraduate pages.