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Title: Characterisation of ADAM15-mediated changes to cellular behaviour in breast cancer cells
Author: Mattern, Jens
ISNI:       0000 0004 6057 8234
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2016
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ADAM15 is a multidomain multifunction transmembrane protein. It participates in protein ectodomain shedding via the metalloproteinase domain. ADAM15 also interacts with integrins via the disintegrin domain. ADAM15 mRNA is subject to complex processing, generating different isoforms as a result of alternative splicing. The splicing affects the intracellular domain (ICD) of the protein, generating distinct SH3 domain binding regions. The expression of specific splice forms correlates with breast cancer prognosis. We aimed to test if ADAM15 isoforms have distinct functions that may affect multiple aspects of cell behaviour in isoform-specific unique ways. We generated an isogenic panel expressing each ADAM15 isoform in MDA-MB-231 acells. Comparative characterisation of the panel demonstrated distinct differences between isoforms, such as catalytic function dependant or independent effects on proliferation rate, changes in cell size, isoform-specific reorganisation of the actin cytoskeleton. We discovered ADAM15 isoform-dependant upregulation of tight junction protein claudin1. Immunofluorescence analysis demonstrated co-localisation of ADAM15 with claudin1 and another tight junction protein ZO1 at cell-cell junctions. Further immunoprecipitation analysis showed potential complex formation with ADAM15 and ZO1. Pharmacological analysis showed claudin1 upregulation is mediated via PI3K/mTOR-pathway. Claudin1 upregulation depended on the metalloproteinase catalytic function of ADAM15, suggesting that ADAM15 isoforms have distinct substrates. Claudin1 was also found in the nucleus of MDA/ADAM15 A expressing cells where it could potentially function as a transcription factor. Focal adhesion (FA) turnover was influenced by ADAM15 isoform expression. In MDA/ADAM15 C expressing cells impairment of FA reassembling was observed, while ADAM15 D expression showed reduced FA disassembly. Comparison of spreading of ADAM15 isoform expressing cells on fibronectin (FN) and vitronectin (VN) revealed that most cell lines preferred FN over VN. Expression of ADAM15 E reduced this preference while ADAM15 A altered it in favour of VN over FN. Changes in subcellular localisation of β3, but not β1-integrin was observed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: RC0254 Neoplasms. Tumors. Oncology (including Cancer)