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neuroscience

Name: Dr. Sarah Newbury

Academic position: Reader in Cell Biology

Research: RNA stability in development

Contact details:

2.08, Medical Research Building
University of Sussex
Falmer
Brighton BN1 9PS
UK

Tel: +44 (0)1273 877874
Fax: +44 (0)1273 877884

E-mail:

Biography:

• BSc (Hons) 1st Class Honours in Biological Sciences, University of Edinburgh. 1976 - 1980.
• PhD in Genetics. Department of Genetics, University of Leeds. 1980-1984.
• Postdoctoral research assistant, Department of Genetics, University of Leeds, 1983 - 1985.
• Postdoctoral research assistant, Department of Biochemistry, University of Dundee, 1985 - 1988.
• Postdoctoral research assistant, Department of Biochemistry, University of Dundee, 1989 - 1989 (part-time).
• Postdoctoral research assistant, Department of Obstetrics and Gynaecology, Ninewells hospital, Dundee, 1989 - 1990 (part-time).
• Royal Society University Research Fellowship 1990- 1998. (part-time from 1990-1993), Initially at Department of Biochemistry, University of Dundee; then from 1/3/91, University of Portsmouth.
• Lecturer, Department of Biochemistry, University of Oxford, 1998 -2004.
• Non-stipendiary College Lecturer at St. Johns College, University of Oxford. 1998 - 2004.
• Lecturer, Institute of Cell and Molecular Biosciences, University of Newcastle-upon-Tyne. 2004 - 2007.
• Senior Lecturer in Cell Biology, Brighton and Sussex Medical School. 2007 - 2009
• Reader in Cell Biology, Brighton and Sussex Medical School. 2009- present.

Teaching focus:

Genetics, cell biology

Research focus:

Development from a single egg to a complex multi-cellular organism requires genes to be switched on and off in particular cells at the correct time. One of the ways in which genes can be switched off is by destroying the messenger RNA, which is the molecule that transmits the information from the DNA in the nucleus to the cytoplasm, where the proteins are made. From work on the yeast, S. cerevisiae, it is known that degradation of RNA is controlled by by ribonucleases and other factors that work together as a “molecular machine”. The research of my group aims to understand the ways in which this novel mechanism of gene regulation can control early development, stem cell differentiation and wound healing in the model organism Drosophila melanogaster.

Current research:

In order to understand the mechanisms controlling stability of mRNAs in the model organisms Drosophila and C. elegans, we are characterising ribonucleases and associated factors and analysing how they interact with each other and with other cellular RNAs and proteins. We have identified nine Drosophila ribonucleases (or associated proteins) and found that all of those characterised are differentially regulated throughout development. This suggests that mRNAs may be degraded via different pathways at particular stages of development and that varying the levels of ribonucleases could specifically regulate gene expression. All of these ribonucleases are extremely well conserved in all eukaryotes suggesting an ancient and conserved mechanisms for the degradation of RNAs.

Ongoing experiments

At present, the work of my group is focussed on the analysis of two proteins that are involved in RNA turnover: a 5’ – 3’ ribonuclease pacman and a 3’-5’ exoribonuclease tazman (dis3). We are using genetic, post-genomic, biochemical and molecular biological techniques to analyse the function of these genes and their role in development.

Role of pacman in epithelial sheet sealing and wound healing

The Drosophila gene pacman is highly homologous to the major yeast exoribonuclease XRN1 and is the only known cytoplasmic 5’ – 3’ exoribonuclease in eukaryotes. We have constructed a number of mutations in pacman by P-element excision and characterised these mutations at the molecular level. Analysis of the 9 viable mutant alleles show that pacman is involved in at least 6 stages of development: female fertility, male fertility, gastrulation, thorax closure, wing development and neural development. In adult flies, the wings are crumpled and sometimes blistered, and in some flies the legs are crooked. Similar defects are found in flies with mutations in cell adhesion molecules, such as integrins and laminins. In flies carrying stronger alleles there are severe defects in thorax closure such that mutants have severe clefts along the back of the thorax. In embryos carrying other alleles, there is a severe defect in gastrulation. Invaginations occur at inappropriate places on the embryo, and the major invaginations can proceed so far that they almost split the embryo in two. These mutant phenotypes suggest that pacman controls some aspect of cell adhesion or changes in cell shape.

In order to determine whether pacman homologues have similar functions in other organisms, we have investigated the effect of the C. elegans homologue xrn-1 on development. In C. elegans, silencing of xrn-1 using RNA interference results in embryos that fail to complete ventral enclosure. xrn1(RNAi) embryos fail to elongate correctly and incomplete enclosure results in cells that are normally internal being bulged out of the embryo. Ventral enclosure in C. elegans is a very similar process to dorsal closure or thorax closure in Drosophila. In both processes, the outer layer of cells stretches and then closes over underlying cells in a “purse-string” movement.

These results therefore reveal an unsuspected link between RNA stability and morphogenetic movements and show, for the first time, that the 5’ – 3’ RNA degradation pathway is crucial at critical stages of development. Morphogenetic movements that occur during dorsal closure and thorax closure in Drosophila and ventral enclosure in C. elegans have been shown to be similar to other cell movements such as wound healing. Indeed, our work in Drosophila also shows that pacman is required for the wound healing process. Therefore, this work has relevance for understanding the processes underlying wound healing in human tissues. To determine whether the human homologue of pacman is up or down-regulated in human cells during wound healing, I have established a collaboration with Professor Tom Lennard (Surgical and Reproductive Sciences, Newcastle).

How can a exoribonuclease specifically affect morphogenesis? In Drosophila, dorsal and thorax closure are known to be controlled by a JNK kinase signalling pathway. A serine/threonine phosphatase, puckered, regulates the JNK kinase (basket). We have recently shown that pacman genetically interacts with puckered suggesting that pacman specifically targets a member of this JNK pathway. This JNK kinase pathway is conserved in human cells and is also known to be involved in wound healing

Role of pacman in neural cells

We have recently shown that pacman protein is localised within cytoplasmic granules in Drosophila neuronal cells. Pacman is co-localised with the decapping protein Dcp1, the Fragile-X mental retardation protein dFMR1 and translation repression factors such as Me31B (Dhh1 in yeast). These granules are likely to be analogous to P-bodies (processing bodies) in yeast, where degradation of mRNAs takes place. Our preliminary evidence suggests that pacman affects dendritic branching and neuromuscular function. These neuronal granules are likely to be critical for local post-transcriptional regulation and may be crucial for dendritic growth and neuronal plasticity.

Interestingly, the pacman protein includes a perfect CAG repeat (CAG) 9 which encodes polyglutamine. Expansions of trinucleotide repeats are known to cause neurodegenerative disease in humans. Our work on the structure of this and other trinucleotide repeats at the RNA level shows that they form a highly stacked triplex-like conformation. We have noticed that pacman protein also includes a repeat (QEAQ) which is found very close to the poly-glutamine repeat. Drosophila huntingtin also includes both the repeats. In addition, polyglutamine repeats are often present in proteins targeted to P-bodies, so at the protein level, these repeats may be important for the integrity of P-bodies. Since some of our mutations in Pacman remove one or both repeats, we have the tools to analyse the function of these repeats, with respect to Pacman activity, in vitro and in-vivo.

Role of pacman in testis cells

Recently, we have shown that Pacman is localised to a small number of discrete foci within the cytoplasm of testis cells undergoing mitosis. In spermatocytes, prior to the meiotic divisions, Pacman is localised in curious cup-shaped structures. Pacman is partially co-localised with Fragile-X mental retardation protein dFMR1 in these particles. We have shown that mutations in pacman affect male fertility and also the size of the testis. We at present investigating the candidate targets of Pacman in testis cells.

dis3

The Drosophila gene dis3 is a 3’-5’ exoribonuclease which is closely related to the E. coli genes RNaseII andRNaseR. In the yeasts S. cerevisiae and S. pombe, mutations in the homologous genes RRP44 and Dis3 lead to defects in mitosis. Both human and S.pombeDIS3 bind, in-vitro, to the GTPase RAN, which is involved in nuclear trafficking, therefore Dis3 may be involved in cellular functions other that RNA stability. Recently, dis3, which is a component of the exosome complex, has been shown to be associated with elongating RNA polymerase, suggesting that is is involved in RNA surveillance during transcription. In collaboration with Dr Cecilia Arraiano, we are characterising the expression of Drosophiladis3 at the protein and RNA level and are also analysing its activity in-vitro. Dr Arraiano’s group are making progress in the structural analysis of RNaseII : since RNaseII and DIS3 are very similar throughout the entire length of the protein, this structural information will shed light on the mechanism of action of dis3 in Drosophila.

 

Key/recent publications:

Barbee, S.A., Estes, P.S., Cziko, A., Luedeman, R.A., Coller, J.M., Johnson, N. Howlett, I.C., Geng, C. Brand, A. Newbury, S.F., Levine, R.B., Wilhelm, J.E., Nakamura, A., Parker, R. and Ramaswami, M. (2006) Staufen and FMRP containing neuronal RNPs are structurally and functionally related to somatic P-bodies. Neuron 52, 997-1009.

Vlad C. Seitan, Peter Banks, Steve Laval, Nazia A. Majid, Dale Dorsett, Amer Rana, Jim Smith, Alex Bateman, Sanja Krpic, Arnd Hostert, Robert A. Rollins, Hediye Erdjument-Bromage, Paul Tempst, Claire Y. Benard, Siegfried Hekimi, Sarah F. Newbury, & Tom Strachan(2006) Metazoan Scc4 homologues link sister chromatid cohesion to cell and axon migration guidance. PLoS Biology 4(8), 1411 – 1425.

Newbury, S.F., Mühlemann, O. and Stoeklin, G. (2006) Turnover in the Alps; an mRNA perspective. EMBO reports 7(2), 143-8.

Newbury, S.F. (2006) Control of mRNA stability in eukaryotes. Biochemical Society Transactions 34, 30-34.

Cairrao, F., Arraiano, C.M. and Newbury, S.F. (2005) The Drosophila gene tazman, an orthologue of the yeast exosome component Rrp44/Dis3, is differentially expressed during development. Developmental Dynamics 232. 733-737.

Black, D. and Newbury, S.F. (2004) RNA interference: a potent gene regulator. The Biochemist 26, 7-10.

Black, D. and Newbury, S.F. (2004) RNAi: How it works and its uses as a technology. The Biochemist 26, 46-47.

Newbury, S. and Woollard, A. (2004) The 5’ – 3’ exoribonuclease xrn-1 is essential for ventral epithelial enclosure during C. elegans embryogenesis. RNA 10, 59-65.

Pinheiro, P. Scarlett, G., Rodger, A., Rodger, P.M. Brown,T., Newbury, S.F. and McClellan, J.A. (2002) Structures of CUG repeats in RNA: Potential implications for human genetic diseases. Journal of Biological Chemistry 277, 35183-35190.

Seago, J.E., Chernukhin, I.V. and Newbury, S.F. (2001) The Drosophila gene twister, an orthologue of the yeast helicase SKI2, is differentially expressed during development. Mechanisms of Development 106, 137-41.

Sparkes AC, Mumford KL, Patel UA, Newbury SF, Crane-Robinson C. (2001) Characterization of an SRY-like gene, DSox14, from Drosophila. Gene. 272,121-9.

Chernukhin, I.V, Seago, J.E and Newbury, S.F. (2001) The Drosophila 5' - 3' exoribonuclease pacman. Methods in Enzymology. 342, 293 – 302.

 

Current/recent laboratory funding:

• BBSRC
• MRC
• Royal Society
• RVI Breast Cancer Research Appeal
• Genetics Society
• Nuffield Foundation

 

Active collaborations:

National:

• Professor Tom Lennard and Dr. Simi Ali, Surgical and Reproductive Sciences, University of Newcastle – Analysis of wound healing in Drosophila and human breast cancer cells
• Professor Tom Strachan, Institute of Human Genetics, University of Newcastle – Analysis of the function of pqn-85, the homologue of the gene for Cornelia-de-Lange syndrome, in C. elegans
• Professor John McCarthy ( University of Manchester) – Biochemical analysis of ribomucleases
• Dr. Simon Bullock, MRC Laboratory of Molecular Biology, Cambridge – Role of a novel gene in spermatogenesis
  

International:

• Dr Cecilia Arraiano ITQB, University of Lisbon, Portugal – Analysis of 3’ – 5’ ribonucleases
• Dr. Tze-Bin Chou, National Taiwan University – Investigation of the function of ribonucleases in oogenesis
  

Other information:

• Member of the Genetics Society
• Member of the British Society for Developmental Biology
• Local ambassador for the the UK Biochemical Society
• Member of the BBSRC “Genes and Development” grant awarding panel