Student Opportunities
We currently have several Honours, Masters or PhD projects available in the Rasko Lab.
Please contact us to express your interest.
Cancer & Gene Regulation Lab
(Supervisors Dr. Chuck Bailey & Prof. John Rasko)
Molecular modeling of cancer-associated mutations in tumour suppressor genes
Description: The ‘master weaver of the genome’ protein CTCF, is mutated in endometrial cancer, as well as colorectal, stomach, breast and haemopoietic cancers. Our group was the first to demonstrate that the ubiquitous zinc finger (ZF) protein CTCF acts as a tumour suppressor gene (Rasko, et al., 2001; Tiffen, et al., 2013). Missense mutations in CTCF are enriched in the DNA-binding ZF region. Our data has showed that mutations in the ZF region can result in a loss-of-function or even gain-of-function in CTCF, which has implications for cancer development.
We are currently modelling ‘hotspot’ mutations on published CTCF ZF protein structures to assess the impact of those mutations. In addition, we will develop homology models for the ZF domain of CTCF-Like protein (CTCFL, also known as BORIS). CTCFL which shares 80% homology with CTCF within its ZF domain, and which is phenotypically similar to CTCF is aberrantly expressed in more than half of all cancers. Both factors have overlapping and unique DNA binding characteristics. We will examine which residues are critical for binding of CTCF and CTCFL to the same DNA target sites and which ZF residues confer DNA binding specificity. This will provide important insight into the sibling rivalry that exists between CTCF and CTCFL in normal biology and cancer.
Students: Honours, Masters, PhD
Skills/Tools: Molecular docking, homology modeling, molecular dynamics simulation, site-directed mutagenesis, molecular cloning, lentiviral gene transfer, mammalian cell culture, chromatin immunoprecipitation.
Understanding the role of CTCF genetic deletion in aggressive endometrial cancer
Description: CTCF is essential for the normal organisation of DNA in cells. Our team has discovered that CTCF is genetically deleted at high rates in the most aggressive and deadly types of endometrial cancer (Marshall, et al., 2017). CTCF deletion predominantly occurs in the Type II serous subtype of endometrial cancer and is associated with poorer overall survival in patients with serous tumours. Additionally we have shown that CTCF deletions also occur in the clear cell subtype and this may be associated with tumour relapse and/or metastasis.
Our culturing of endometrial cancer cell-lines as 3D spheroids has shown that a functional consequence of CTCF deletion in results in a loss of cell polarity – an early event in endometrial cancer pathology. Our analysis of gene expression data in CTCF heterozygous endometrial tumours has revealed a widespread dysreguation of transcription. In this project we will those examine genes and biochemical pathways that are dysregulated in CTCF mutant endometrial cancers. We will investigate the impact of these genes on cell polarity in 3D spheroids which will give us important insights into early pathophysiological events underlying endometrial cancer.
Students: Honours, Masters, PhD
Skills/Tools: Mammalian cell culture, spheroid culture, retroviral gene transfer, qRT-PCR, cell biology assays, shRNA knockdown, CRISPR/Cas9 gene editing, flow cytometry, Western blotting, mouse work (Ctcf+/- mice), confocal microscopy.
Modulation of host entry factors to improve AAV-mediated gene therapy
Description: Recombinant adeno-associated virus (rAAV) has gained widespread use as a gene delivery vector for corrective gene therapies due to its lack of association with any human disease and its ability to safely and efficiently deliver a genetic payload into a broad range of tissues. For liver-specific genetic diseases, current rAAV modalities have not provided the necessary high-level transduction efficiencies and humoral neutralisation properties necessary for curative outcomes in diverse patient groups.
Recent efforts have improved the transduction efficiency of rAAV vectors by engineering capsids with higher affinity or cell-specific tropism and increased resistance to neutralising antibodies. Indeed, a third avenue for increasing AAV-mediated therapeutic efficacy, modulating host entry factors to increase AAV capsid entry, remains unexplored. KIAA0319L was recently shown to be an essential host entry factor for most AAV serotypes, however the biology and normal function of KIAA0319L is poorly understood. In this project, we will use a combination of biochemical, genetic and proteomic strategies to functionally characterise the host determinants that regulate KIAA0319L expression and distribution. The overall goal is to test the hypothesis that modulation of host entry factor expression can improve gene transfer efficiency in clinically relevant circumstances.
Topics that may be explored in this research project include:
1. Mutagenesis studies of KIAA0319L to identify motifs important for trafficking
2. Perform mass spectrometry to identify binding partners that regulate KIAA0319L localisation and function
3. Identify post-translational modifications that regulate KIAA0319L localisation
4. Examine KIAA0319L spatiotemporal distribution during AAV transduction using confocal microscopy
Students: Honours, Masters, PhD
Skills/Tools: Mammalian cell culture, retroviral gene transfer, cell biology assays, CRISPR/Cas9 gene editing, mass spectrometry, confocal microscopy, flow cytometry, Western blotting, qRT-PCR.
The role of MGA mutation in chronic lymphocytic leukaemia
Description: Chronic lymphocytic leukaemia (CLL) is the most common leukaemia in senior Australians. Every year nearly 1000 Australians are diagnosed with CLL and typically 80% of all new diagnoses are in patients over the age of 60 years. CLL is a slow developing cancer affecting B cells. Genetic mutations acquired in these B cells result in their transformation into cancerous cells that can live longer and grow faster than normal B cells. Similar to many blood cancers, genetic alterations in CLL can be heterogeneous, and include point mutations, chromosomal deletions, amplifications and rearrangements. Recent reports have identified the gene encoding the transcription factor Max Gene Associated (MGA) to be recurrently deleted in CLL. MGA inactivation through chromosomal deletion (del15q15) or point mutation occurs in 4% of CLL, but this increases to 16% as CLL disease progresses to chemotherapy resistance.
Our hypothesis is that genetic inactivation of MGA promotes chronic lymphocytic leukaemia disease progression. We will test this hypothesis by analysing how acquired genetic lesions in MGA alter the proliferation, differentiation and survival of CLL cells and contribute to cellular transformation.
Topics that may be explored in this research project include:
1. Determining the consequences of MGA mutations using in vitro functional assays.
2. Structure/function correlations of mutations in MGA using homology modelling.
3. CRISPR/Cas9-induced inactivation of MGA alleles in CLL cells
4. Analysis of haemopoietic cell populations in Mga+/- mice.
Students: Honours, Masters, PhD
Skills/Tools: Cloning, cell biology assays, retroviral gene transfer, flow cytometry, Western blotting, immunofluorescence, immunoprecipitation, CRISPR/Cas9, RT-qPCR, next generation sequencing, mouse handling.
Li Ka Shing Cell & Gene Therapy Program
(Supervisors Dr. Dannel Yeo & Prof. John Rasko)
Circulating Tumour Cells to Monitor Cancer Patients
Description: Circulating tumour cells (CTCs) are tumour cells that have been released from the primary tumour tissue to form metastases by travelling through the blood and lymphatic system. Capturing and analysing these rare cells is now possible using our next generation liquid biopsy platform. We are able to identify these CTCs. Hence, this platform has the potential to provide ‘real-time’ cancer monitoring throughout all stages of a patient’s cancer journey and identify potentially effective treatments in deadly cancers such as pancreatic cancer, lung cancer, and mesothelioma. Potential research topics that can be incorporated into an honours project include:
1. Characterising CTCs using genetic, cellular and imaging techniques;
2. Evaluating the ability of circulating tumour cells to predict patient response;
3. Establishing and characterising patient-derived CTC organoid cultures;
4. Evaluating CTCs and other blood biomarkers as a diagnostic marker to improve early detection.
Students: Honours, Masters, PhD
Skills/Tools: Mammalian cell culture (3D), cell biology assays, western blot, RT-qPCR, microscopy, immunofluorescence, cell picking, mouse models.
Investigating CAR-T Therapy for Pancreatic Cancer
Description: Chimeric antigen receptor-engineered T-cell (CAR-T) therapy is an exciting new cellular immunotherapy for the treatment of cancer. CAR-T therapy are now approved for blood cancers but the same success has not been observed in solid cancers. Isolated patient T-cells are modified to target a specific tumour surface antigen and then injected back into the patient. Novel CAR-T therapies will be evaluated in pancreatic cancer using the latest cancer model systems including 2D cells, 3D organoids and mice models. To test the efficacy of CAR-T cells, cytotoxicity and immune activation/persistence will be evaluated, and mechanisms of resistance will be explored.
Students: Honours, Masters, PhD
Skills/Tools: mammalian cell culture, cell biology assays (including cellular impedance assays, live cell imaging), isolating T cells, lentivirus gene transfer, immune phenotyping (by flow cytometry), RT-qPCR, cytokine analysis (ELISA), and mouse models.
Computational BioMedicine Lab
(Supervisor A/Prof. Ulf Schmitz)
Intron retention-mediated microRNA sponging in cancer
Description: In a phylogenetic study of intron retention in vertebrate species we have identified a potential role of retained introns to act as conserved microRNA (miRNA) sponges that are involved in the fine-tuning of global gene expression regulation (Schmitz et al., 2017). miRNAs have dozens to hundreds of targets and intron retention (IR) can affect hundreds or even thousands of expressed genes, depending on cell/tissue type and condition. Both miRNA-induced translation repression or target degradation and IR-induced nonsense-mediated decay are concurrent mechanisms of gene expression control. In addition to that, we have predicted a density of >100 miRNA response elements per kilobase in retained introns (in human granulocytes). It is therefore important to analyse concurrent and competitive gene expression regulation including the effects of IR/miRNA sponging on a global level.
In this project we will identify gene-regulatory network modules using a data integration approach to determine patterns of competitive post-transcriptional gene regulation. Toward this, we will integrate expression profiles of miRNAs and genes, IR pattern and predicted miRNA–gene interactions, TF–gene interactions, and IR-miRNA sponge interactions. For a selected sub network we will construct a mathematical model of competitive post-transcriptional gene regulation. We will parameterise this model using data from in vitro time series experiments in which we perturb the identified network module. We will use our model to simulate the effects of increased/reduced IR abundance in cancer cells to study system dynamics and the effect of varying IR levels on the concentration of endogenous miRNA targets. We will also determine whether our model can be adopted and might help to explain aberrant gene expression patterns in cancer cell lines.
Students: Honours, Masters, PhD
Skills/Tools: Bioinformatics data integration, systems biology, COPASI, PCR, ELISA, gene/miRNA knock down (shRNA, antagomiR), plasmid design.
Development of an Intron Retention database
Description:In a large-scale analysis of alternative splicing across 2,500 human tissue samples and cell lines we generated a wealth of data regarding gene-, cell type-, tissue-, and disease-specific intron-retention events (Middleton et al., 2017). This data is in parts accessible through a rudimentary web interface: here
In this project we will develop a sophisticated database and web interface design to provide an efficient and rich user experience facilitating a rapid success in the hunt for information about intron-retention. The new IRBase 2.0 will provide data in interactive graphs and customized data retrieval options.
Students: Honours, Masters
Skills/Tools: HTML, JavaScript, R Markdown (Shiny, reveal.js), MySQL, CGI, CSS.
Cooperating microRNAs for cancer therapy - a systems medicine approach
Description: Drug resistance remains one of the most significant problems in cancer therapy and requires a systematic therapeutic approach. In this project, we use an established systems biology approach to design a therapeutic strategy to tackle chemoresistance in aggressive tumours (Lai et al., 2018). We will utilize pairs of cooperating microRNAs as co-adjuvants to sensitize tumour cells to existing anti-cancer drugs.
We will use secondary structure prediction, 3D docking and molecular dynamics simulations as well as kinetic modelling to identify cooperating microRNA pairs and their synergistic potential in regulating drivers of chemoresistance. We will validate our predictions experimentally using adenoviral vector-based microRNA transfection, qRT-qPCR, luciferase assays, western blotting and/or ELISA assays. We will test this strategy by applying a genotoxic drug and a pair of cooperating microRNAs that repress the expression of chemoresistance-genes in order to overcome resistance in aggressive tumour cell populations.
Students: Honours, Masters, PhD
Skills/Tools: Bioinformatics, structural biology, systems biology, network modelling (graph theory), CellDesigner, Cytoscape, Copasi, molecular biology assays (adenoviral vector-based microRNA transfection, qRT-qPCR, luciferase assays, immunoblotting and/or ELISA assay.
Deciphering the role of introns in cancer
Description: Discontinuous gene structure is one of the hallmarks of eukaryotic genomes. Protein-coding sequences are intervened by ‘introns’, allowing a greater shuffling of the coding sequences that increases diversity of the proteins produced from a given locus. Most introns were thought to play a passive role in gene regulation as they are usually removed from the transcribed RNA during RNA maturation. However, several seminal studies from our lab discovered that a significantly greater number of genes exhibit a phenomenon known as ‘intron retention’ (IR). Our lab discovered that RNA bearing IR exhibited a higher rate of termination codons because of presence of the retained intron, leading to degradation of the transcript by a process known as non-sense mediated decay and ultimately resulting in a reduction in gene expression (Wong et al (2013), Cell).
Our lab aims to investigate the role of IR in cancer. We have measured levels of roughly 200,000 introns using RNA-sequencing in multiple types of cancer. In this project, the student will apply algorithms and test computer predictions with wet lab experiments to confirm IR events in human cancers. Specifically, the student will be testing whether an IR event is detected in cells using RT-qPCR, identify its ‘signature’ in RNA-sequencing data and modify our detection algorithm accordingly to enhance the discovery of biologically important IR events. This project will help prioritise a set of IR events and their potential role in cancer biology.
Students: Honours, Masters, PhD
Skills/Tools: Cell culture, RNA extraction, primer design, RT-qPCR and exposure to RNA-sequencing/bioinformatics. Knowledge of the programming language R will be advantageous.
Characterizing fusion transcripts in cancer and normal cells
Description: DNA rearrangement leading to fusion genes is a hallmark of cancer. A number of these fusions are used as biomarkers and therapeutic targets in different cancer types; prominent examples include BCR-ABL1 for chronic myeloid leukemia and MYC-IGH for Burkitt's lymphoma. Traditionally, DNA rearrangement and gene fusion was attributed largely to chromosomal translocations. However, recent years have produced conclusive evidences that two or more transcripts can fuse at the transcription level irrespective of chromosomal translocations.
In this project, we will analyse long-read sequencing and single-cell sequencing data from tumour samples and healthy tissue to identify known and novel fusion transcripts and determine their (cancer) cell-type specific expression profiles.
Students: Honours, Masters
Skills/Tools:
Mirtron synthesis and expression in leukemia
Description: In a large-scale analysis of alternative splicing across 2,500 human tissue samples and cell lines we generated a wealth of data regarding gene-, cell type-, tissue-, and disease-specific intron-retention events . This data is in parts accessible through a rudimentary web interface: http://mimirna.centenary.org.au/irfinder/database
In this project we will develop a sophisticated database and web interface design to provide an efficient and rich user experience facilitating a rapid success in the hunt for information about intron retention. The new IRBase 2.0 will provide data in interactive graphs and customized data retrieval options.
Students: Honours, Masters
Skills/Tools:
Core transcriptional networks in cell trans-differentiation
Description: Core transcriptional networks are essential drivers and determinants of cell-fate transitions. To date, our mechanistic understanding of these essential regulatory layers is very limited, especially in the context of trans-differentiation, despite being one of the most promising therapeutic cell replacement strategies in regenerative medicine. In this project, we will reconstruct gene-regulatory networks involving non-coding RNAs and mRNAs that drive trans-differentiation of human cells and identify key alternative splicing events during cell-fate transitions.
Students: Honours, Masters, PhD
Skills/Tools: