Research Project 1: Self-Assessment

Piece of work being assessed:

Research project report and poster presentation.

Title:

Investigating the human ZDHHC9 protein-protein interaction network.

Description:

10-week[] long laboratory project using a new method to screen for protein-protein interactions
(PPIs), under the supervision of Prof Chris Sanderson. The project has been assessed via: a research
report, written in the style of an academic journal; a poster presentation; and a mini viva session with
the external assessor of the report, Dr Sudeep Pushpakom. Based on a previously
performed[published?] PhD thesis in the lab[?], this project investigated potential interaction partners
of the membrane protein ZDHHC9, while using the membrane yeast two-hybrid (MYTH) assay to
screen a cDNA library of membrane proteins. ZDHHC9 is a palmitoyltransferase, known to cause X-
linked mental retardation, and is also involved in the development of cancer through interactions with
HRAS and NRAS proteins.
Aims:

This project aimed to build a novel network of potential interactors of the ZDHHC9 protein, all while
using the MYTH screen confidently. Therefore[?], I aimed to test the utility of a newly-optimised Gap
repair technique to use as a high throughput method for the PPI screens, as the Gap repair step is an
important step in confirming results obtained through the MYTH screen. I achieved writing a high-
quality scientific report, and presented my results in a poster presentation in a clear and structured
way.
Learning Objectives: [consider past tense?]

a) Performing the MYTH screen unassisted and with confidence.

b) Analysing the gained results using databases like NCBI Nucleotide blast, uniprot.org, etc.
c) Presenting my results in a comprehensive research report, poster presentation, and mini viva.
d) Being critical with the obtained results, and reading literature to give myself an overview of
the current developments in the field.
e) Improving organisational and communication skills.
Achievements:

I learned how to use the MYTH assay confidently by myself, which was a completely new technique for
me. I improved my understanding of the relevance of interaction networks as a basic research method,
and learned more about the importance of palmitoylation processes in general. I enhanced my ability
to work in stressful situations, all the while learning a new technique, understanding it, and then using
it to produce high-quality results of publication standard[]; and all in 10 weeks[needed?]. Through
journal clubs, group meetings, and general close interaction with other members of the laboratory and
my supervisor I came to improve my scientific organisation skills.
Learning Outcomes:

a) Improving my scientific writing skills and presenting results confidently.

b) Adding a new technique to my set of laboratory skills.
 Research Project 2: Self-Assessment

Piece of work being assessed:

Research project report and poster presentation.

Title:

Investigating novel ZDHHC9 interaction partners.

Description:

10-week[] long laboratory project using new methods to confirm a previously established protein-
protein interaction (PPI) network under the supervision of Prof Chris Sanderson. The project has been
assessed via: a research report, written in the style of an academic journal; a 12-min Power Point
presentation; and a mini viva session with the external assessor of the report, Dr Ian Copple. Based on
a previously performed PhD thesis[see above] in the lab, this project investigated potential interaction
partners of the membrane protein ZDHHC9, using PCR amplification, Gateway cloning technique,
western blotting, and confocal live-cell imaging. ZDHHC9 is a palmitoyltransferase, known to cause X-
linked mental retardation, and is also involved in cancer disease development[see above] through
interactions with HRAS and NRAS proteins.
Aims:

This projected intended to confirm putative novel interactions partners of the ZDHHC9 protein by
using a set of novel techniques. I planned to confirm HRAS and NRAS interactions with the protein of
interest, ZDHHC9, as described in the literature, and, further on, to confirm novel putative interaction
partners in mammalian cells. As a basic criterion[i.e requirement?] of investigating PPIs, I aimed to
perform basic microscopy analysis of co-localisations of proteins, including analysis of potential FRET
signals.
Learning Objectives:

a) Designing of primers for PCR amplification, according to Guidelines for Gateway primers.
b) Performing Gateway cloning technique, basic confocal live-cell imaging
c) Improving my ability to write a scientific report more easily and fluently
d) Being confident using databases and finding up-to-date literature

Achievements:

In this technically difficult project, I learned to use a set of novel techniques autonomously, which was
very challenging in this short time period. I developed a better understanding of the processes of
techniques in the laboratory used/and used them[?] to confirm previous results obtained through
other techniques. As a non-native speaker, I improved my scientific writing and communication skills
rapidly in this rotation. I also learned to perform the preparation steps of the single techniques, while
designing Gateway primers by myself. I was able to present my whole project confidently and clearly in
an oral presentation.
Learning Outcomes:

a) Working even more independently

b) Presenting lots of results in a well-structured oral presentation
 Research Project 3: Self-Assessment

Piece of work being assessed:

Research project report and poster presentation.

Title:

Secondary investigation of proteins involved in hereditary spastic paraplegia (HSP).

Description:

This project intended to investigate protein-protein interactions (PPIs) of proteins involved in a group
of inherited motor neuron disorders, called hereditary spastic paraplegia (HSP). The project was
restricted to 10 weeks of laboratory work under the supervision of Prof Chris Sanderson. Afterwards,
the project was assessed via: a report, written in the style of a scientific journal; a poster presentation;
and a mini viva session with the external marker of the report, Dr Dean Naisbitt. Based on a previously
performed PhD thesis in the lab about several autosomal-recessive inherited HSP genes, we decided to
further investigate some of the strong interaction partners[] using Gateway cloning (to produce
fluorescent expression vectors), western blotting, confocal live-cell imaging, and pull-down analysis.
Aims:

The aims of this project included becoming even more confident with the already performed
techniques used to investigate the previously[?] via yeast two-hybrid screens established putative
interaction network in mammalian cells.[*brain implodes*] Therefore, it was important to focus on
molecular techniques and intensify microscopy-based analysis of interactions and co-localisation of the
proteins. I aimed to learn a novel technique, Pull-down analysis, as a method for purification affinity
screens of PPIs.
Learning Objectives:

a) Improving skills for confocal microscopy-based analysis to a level that requires very little
outside assistance
b) Learning how to perform pull-down analysis
Achievements:

After this challenging rotation, I am nearly able to perform confocal live-cell imaging by myself, and
have greatly gained confidence in analysing interaction signals of FRET analysis or bleaching
experiments. As the last project was not as successful as the ones before it, I had to learn how to
handle stressful and disappointing situations. I was able to complete pull-down analysis by myself and
gain results of good quality within this short period of time.
Learning Outcomes:

a) Handling stressful and frustrating situations in a more positive way

b) Improving skills related to creating a clear and well-structured poster outline