Investigation of p75 neurotrophin receptor and the role of its post-translational modifications in tumour cell motility and invasiveness

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dc.check.date10000-01-01
dc.check.embargoformatBoth hard copy thesis and e-thesisen
dc.check.entireThesisEntire Thesis Restricted
dc.check.infoIndefiniteen
dc.check.opt-outNoen
dc.check.reasonThis thesis is due for publication or the author is actively seeking to publish this materialen
dc.contributor.advisorMcCarthy, Justin V.en
dc.contributor.authorBerghoff, Janina
dc.contributor.funderHealth Research Boarden
dc.date.accessioned2014-05-12T14:29:36Z
dc.date.issued2013
dc.date.submitted2013
dc.description.abstractThe p75 neurotrophin receptor (p75NTR) is a member of the tumour necrosis factor superfamily, which relies on the recruitment of cytosolic protein partners - including the TNF receptor associated factor 6 (TRAF6) E3 ubiquitin ligase - to produce cellular responses such as apoptosis, survival, and inhibition of neurite outgrowth. Recently,p75NTR was also shown to undergo γ-secretase-mediated regulated intramembrane proteolysis, and the receptor ICD was found to migrate to the nucleus where it regulates gene transcription. Moreover, γ-secretase-mediated proteolysis was shown to be involved in glioblastoma cell migration and invasion. In this study we report that TRAF6-mediated K63-linked polyubiquitination at multiple or alternative lysine residues influences p75NTR-ICD stability in vitro. In addition, we found that TRAF6-mediated ubiquitination of p75NTR is not influenced by inhibition of dynamin. Moreover, we report beta-transducin repeats-containing protein (β-TrCP) as a novel E3- ligase that ubiquitinates p75NTR, which is independent of serine phosphorylation of the p75NTR destruction motif. In contrast to its influence on other substrates, co-expression of β-TrCP did not reduce p75NTR stability. We created U87-MG glioblastoma cell lines stably expressing wild type, γ-secretaseresistant and constitutively cleaved receptor, as well as the ICD-stabilized mutant K301R. Interestingly, only wild-type p75NTR induces increased glioblastoma cell migration, which could be reversed by application of γ-secretase inhibitor. Microarray and qRT-PCR analysis of mRNA transcripts in these cell lines yielded several promising genes that might be involved in glioblastoma cell migration and invasion, such as cadherin 11 and matrix metalloproteinase 12. Analysis of potential transcription factor binding sites revealed that transcription of these genes might be regulated by well known p75NTR signalling cascades such as NF-κB or JNK signalling, which are independent of γ-secretase-mediated cleavage of the receptor. In contrast, while p75NTR overexpression was confirmed in melanoma cell lines and a patient sample of melanoma metastasis to the brain, inhibition of γ-secretase did not influence melanoma cell migration. Collectively, this study provides several avenues to better understand the physiological importance of posttranslational modifications of p75NTR and the significance of the receptor in glioblastoma cell migration and invasion.en
dc.description.statusNot peer revieweden
dc.description.versionAccepted Version
dc.format.mimetypeapplication/pdfen
dc.identifier.citationBerghoff, J. 2013. Investigation of p75 neurotrophin receptor and the role of its post-translational modifications in tumour cell motility and invasiveness. PhD Thesis, University College Cork.en
dc.identifier.endpage288
dc.identifier.urihttps://hdl.handle.net/10468/1555
dc.language.isoenen
dc.publisherUniversity College Corken
dc.rights© 2013, Janina Berghoff.en
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/en
dc.subjectCancer Biologyen
dc.subjectp75 neurotrophin receptoren
dc.subjectUbiquitinationen
dc.subjectGlioblastomaen
dc.subjectGamma-secretase-mediated proteolysisen
dc.subjectMicroarrayen
dc.subjectCancer cell migrationen
dc.thesis.opt-outfalse
dc.titleInvestigation of p75 neurotrophin receptor and the role of its post-translational modifications in tumour cell motility and invasivenessen
dc.typeDoctoral thesisen
dc.type.qualificationlevelDoctoral Degree (Structured)en
dc.type.qualificationnamePhD (Science)en
ucc.workflow.supervisorjv.mccarthy@ucc.ie
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