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Universidade Nacional da Irlanda em Galway -- Ciência sem Fronteiras
PhD Project Template
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Use one form per project
Please complete & submit to international@nuigalway.ie as soon as possible, and by 27/11/2012
In your email, begin the subject line with [SWB] (be sure to use square brackets) to ensure that
your email is filed correctly. Emails will be automatically filed
PI name & contact details:
Dr. Heinz Peter Nasheuer,
Biochemistry,
National University of Ireland, Galway
University Road
Galway, Ireland
Email: h.nasheuer@nuigalway.ie
phone: +353-91-49 2430
fax: +353-91-49 5504
School:
School of Natural Sciences
Has project been agreed with head (or
nominee) of proposed registration school?
Research Centre / group affiliation:
Research group / centre website:
PI website / link to CV:
Centre for Chromosome Biology;
Cell Cycle Control Group
http://www.chromosome.ie/researchers/nasheuer/
http://www.chromosome.ie/researchers/nasheuer/
Brief summary of PI research / research group / centre activity (2 or 3 lines max):
The research group has three major topics of interest:
 Cell cycle-dependent regulation and mechanisms of human DNA replication and repair pathways
 DNA replication of human polyoma viruses defining small molecules and novel small cellular
non-coding RNA as inhibitors of viral DNA replication
 Systems biology of protein-protein interactions in living cells using advanced microscopy
techniques
Title & brief description of PhD project (suitable for publication on web):
P54nrb a novel player in the DNA damage response
Cellular DNA is continuously challenged by various environmental and cellular stresses that damage
DNA. Therefore, mechanisms to maintain genome integrity are important for cell viability and
survival after DNA damage as well as the prevention of inherited diseases in humans such as cancer.
Investigating the protein networks controlling the DNA damage response including DNA repair
pathways has been a focus of biomedical research and cancer cell biology. An important step is to
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Universidade Nacional da Irlanda em Galway -- Ciência sem Fronteiras
understand the recognition of damaged DNA and the establishment of the signal transduction in the
context of chromatin. A screen for human factors binding to chromatin after UV treatment of DNA
reproducibly showed that the protein p54nrb, which is also called nono, increased its binding to
chromatin after UV treatment. p54nrb is an ubiquitous protein that forms a complex with PSF
(Polypyrimidine tract-binding protein-associated splicing factor). The p54nrb-PSF complex has multifunctional characteristics and is involved in nuclear RNA processing, and regulation of transcription.
Our results showed that down-regulation of p54nrb with specific siRNA increased the survival of
these cells after UV treatment. A specific feature of p54nrb after UV treatment is its localization to
nucleoli and this localization depends on the cooperative activity of the two DNA damage response
initiator kinases ATM and ATR. Using full length p54nrb and deletion mutants thereof fused to green
fluorescence protein we determined that the region of p54nrb containing a helix-turn-helix motif is
necessary and sufficient for the localization of p54nrb to nucleoli after UV damage.
In this project, the role of p54nrb in the DNA damage response and factors controlling its localization
to nucleoli will be analyzed. To investigate the localization of p54nrb after UV damage, the region
containing the helix-turn-helix motif will be mutated and the mutant proteins will be expressed in
human cells as fusion protein with green fluorescent protein. After UV treatment, their localization
will be determined using fluorescence microscopy. To investigate the regions and residues
controlling the viability of cells after UV treatment endogenous p54nrb will be down-regulated with
siRNAs and then wild-type p54nrb, deletion mutants, and p54nrb with specific residues exchanged to
alanine will be encoded as cDNAs resistant to siRNA treatment and will be expressed in human cells
(knock-down and knock-in experiment). These cells will then be treated with UV or mock-treated
and the viability of mutant and wild type p54nrb expressing cells will be determined and compared.
Cells expressing mutant p54nrb, which have the same viability as siRNA-treated control cells, have
lost their ability to negatively influence viability of cells after UV treatment. The networks of protein
binding to mutant and wild-type p54nrb will be compared to determine the regulatory elements in
this process. To this end, these mutant proteins and wild-type p54nrb will be affinity-purified and the
co-purifying proteins including PSF will be analyzed using mass spectrometry and verified by
western blotting. These results will provide new functions to protect cells. Moreover, known DNA
damage signaling proteins such as replication protein A, claspin, TopBP1 and ATR will be analyzed
for their phosphorylation status, protein-protein interactions and biochemical activity in these
mutant p54nrb expressing cells.
Unique selling points of PhD project in NUI Galway:
The student will gain experience in multiple areas of genome stability learning state-of-the-art
biochemical, cell and molecular biological techniques including mass spectrometry, siRNA
methologies, advanced fluorescence and immunofluorescence microscopy.
The project leader is a member of the Centre for Chromosome Biology (CCB) at NUI Galway in
Ireland and very closely cooperates with research groups at NUI Galway and with international
laboratories to advance the knowledge in the mechanisms of genome instability. CCB comprises a
highly interactive network of researchers who are interested in topics focusing the biology of
genomes.
The Centre currently consist of 11 groups with more than 90 researchers and support staff
[http://www.chromosome.ie/]. Their interests are focused on all molecular aspects underlying
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Universidade Nacional da Irlanda em Galway -- Ciência sem Fronteiras
chromosome replication, segregation, repair, evolution, chromatin regulation and transcription. To
investigate chromosome biology processes involved the Centre has available a range of technologies
available at the molecular, cellular and organismal level.
The Centre has gained international attention through its recent findings on the regulation of
trinucleotide repeats and the involvement of small noncoding RNA in the inhibition of viral DNA
replication.
Name & contact details for project queries, if different from PI named above:
as above
Please indicate the graduates of which disciplines that should apply:
Graduates with a background molecular gentics/biology, biochemistry, cell biology, or related
disciplines should apply.
Ciência sem Fronteiras / Science Without Borders Priority Area:
Please indicate the specific programme priority area under which the proposed PhD project fits- choose only
one (tick box):
Engineering and other technological areas
Pure and Natural Sciences (e.g. mathematics, physics, chemistry)/Physical Sciences (Mathematics,
Physics, Chemistry, Biology and Geosciences)
Health and Biomedical Sciences / Clinical, Pré-clinical and Health Sciences
X
Information and Communication Technologies (ICTs), Computing
Aerospace
Pharmaceuticals
Sustainable Agricultural Production
Oil, Gas and Coal
Renewable Energy
Minerals, Minerals Technology
Biotechnology
Nanotechnology and New Materials
Technologies for Prevention and Mitigation of Natural Disasters
Bioprospecting and Biodiversity
Marine Sciences
Creative Industry
New technologies in constructive engineering
Please indicate which of the following applies to this project (referring to Science Without Borders
arrangements):
Suitable only as a Full PhD (Y/N):
Available to candidates seeking a Sandwich PhD arrangement (Y/N): _____
Suitable for either:
Y