PhD Studentship: Dissecting Extracellular Matrix Internalisation Mechanisms using Functional Genomics

University of Sheffield - Department of Biomedical Science, The University of Sheffield / Institute of Molecular and Cell Biology, A*STAR institute, Singapore

This is a 4-year funded PhD project, with 2 years in Sheffield and 2 years in Singapore.

The extracellular matrix (ECM) is a complex network of secreted glycoproteins providing tissue support and controls a variety of cell functions, including tumour growth. Reports dating back to the 1990s have documented the internalisation of ECM components, including collagen and laminin. More recently, ECM endocytosis has been linked to increased matrix degradation by cancer cells. Consistent with this observation, previous work by the Rainero group demonstrated that ECM internalisation is required for cancer cell migration and nutrient signalling, suggesting the intriguing hypothesis that ECM uptake might represent a nutrient source for invasive cancer cells. These observations point to the machinery controlling ECM endocytosis as a novel target for the development of pharmacological intervention to limit cancer spreading.

Preliminary data from the Rainero lab show that ECM internalisation is strongly increased in invasive breast cancer cells, compared to normal mammary epithelial cells and non-invasive breast cancer cells. Cells interact with the ECM through plasma membrane receptors, which has been shown to promote the internalisation of their ECM component ligands. The molecular mechanisms controlling this are poorly defined. To faithfully recapitulate the architecture and composition of the in vivo ECM, cell-derived matrices (CDMs) will be used. These are fibrillar 3D matrices generated by fibroblasts and the tools to study their internalisation have been recently developed in the Rainero lab.

This project will characterise the endocytic mechanisms controlling ECM internalisation using a functional genomic approach and test whether the GALA pathway affects ECM internalisation. This will be achieved using the RNA interference (RNAi) screening technology at the genomic scale, which will allow the identification and accurate quantification of novel key players in this process. The RNAi screen will be performed at the Institute of Molecular and Cell Biology RNAi screening facility (which is embedded in the Bard lab), using the human genome siRNA library and the invasive human breast cancer cell line MDA-MB-231. We will use bioinformatics to construct regulatory networks and subnetworks, to provide a genetic overview of the endocytic pathways controlling ECM internalisation by cancer cells. Based on this analysis, key pathway(s) will be identified and a list of hits will be selected and validated using cells extracted from mouse primary breast tumours. Finally, the contribution of the identified regulators of ECM uptake and the GALA pathway in controlling breast cancer cell proliferation and migration will be investigated, using imaging-based proliferation assays, live cell time-lapse microscopy and 3D invasion assays.

These studies will provide a comprehensive and complementary set of molecular cell biology and genetic approaches, coupled with advanced bioinformatics and imaging techniques to elucidate the molecular mechanisms controlling ECM internalisation. The data generated will provide novel insight into the contribution of regulators of ECM internalisation in the control breast cancer cell proliferation and migration.

Whilst in Sheffield, students receive fees (£4,260 in 2018/19) and an RCUK rate stipend (£14,553 in 2018/19).

Whilst in Singapore, students receive the following:

  • A monthly stipend of 2,500 Singapore dollars.
  • A one-off "settling-in allowance" of 1,000 Singapore dollars.
  • A one-time airfare allowance of 1,500 Singapore dollars.
  • Consumables and Bench Fees.
  • Cost of medical insurance while the student is based at A*Star.

For further information please contact E.Rainero@sheffield.ac.uk.

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