Dr Carol Murphy BSc, PhD

Dr Carol Murphy

School of Biosciences
Senior Lecturer in Advanced Biological Imaging

Contact details

Address
School of Biosciences
University of Birmingham
Edgbaston
Birmingham
B15 2TT
UK

Carol is a Senior Lecturer in Advanced Biological Imaging in the School of Biosciences. Her main interest focuses on the role of the endocytic pathway in TGF-β/Activin receptor signalling due to their involvement in angiogenesis, carcinogenesis and development. She is especially interested in understanding the role of endocytic trafficking during pluripotency and differentiation and therefore uses human embryonic stem cells, human induced pluripotent stem cells and mature primary endothelial cells. 

Qualifications

  • BSc (University College Dublin, Ireland)
  • PhD (University College Dublin, Ireland)

Biography

Carol Murphy has a BSc  and PhD from University College Dublin, Ireland. She worked with Ulrich Rüther as a post-doctoral fellow at the Differentiation Programme at the European Molecular Biology Laboratory (EMBL) and also in the Cell Biology Programme at the EMBL in the laboratory of Marino Zerial, Heidelberg, Germany. She then moved to the Laboratory of Biological Chemistry at the University of Ioannina (UOI) Medical School, Greece, where she was funded by the EU as “Experienced Scientist”. Since 2001 she has been working as Researcher B at IMBB-BR. Her research interests include the intracellular trafficking of receptors and the control of signalling with extension to human stem cells and human induced pluripotent stem cells. In July 2015 she moved to the School of Biosciences, University of Birmingham as Senior Lecturer in Advanced Biological Imaging.

Postgraduate supervision

PhD and Masters projects are offered in the general area of membrane trafficking and signalling. Currently our main interest focuses on the role of the endocytic pathway in TGF-β/Activin receptor signalling due to their involvement in angiogenesis, carcinogenesis and development. We are especially interested in understanding the role of endocytic trafficking during pluripotency and differentiation and therefore use human embryonic stem cells, human induced pluripotent stem cells and mature primary endothelial cells. Technical approaches include confocal microscopy, molecular biology and cell culture.

PhD opportunities

Research

Signalling and endocytic trafficking of TGFβ family ligands/receptors

The transforming growth factor β (TGFβ) family of ligands consists of evolutionary conserved pleiotropic secreted cytokines, which include TGFβ1, Activins and bone morphogenetic proteins (BMPs). TGFβ ligands trigger heteromeric complex formation between specific transmembrane type I and type II Ser/Thr kinase receptors, in which the type II receptor transphosphorylates and activates the type I receptor. R-SMADs are phosphorylated by type I receptor, and in turn can form heteromeric complexes with SMAD4. These activated SMAD complexes accumulate in the nucleus, where they directly or indirectly bind to specific promoter region on target genes together with transcription factor and/or co-activators/repressors (Reviewed in Schmierer, B., and Hill, C.S, Nat Rev Mol Cell Biol 2007, 8: 970-982).

 Individual members of this family play crucial roles in multiple processes throughout development and in the maintenance of tissue homeostasis in adult life. As a consequence, deranged signalling by TGFβ family members has been implicated in many human diseases, including cancer, fibrosis, auto-immune and vascular diseases. Indeed, TGFβ signalling plays critical and dual role in the progression of human cancer. At early stages, TGFβ functions as a tumour suppressor, but depending on the context, TGFβ may support tumour progression by enhancing invasion, dissemination and immune evasion (Meulmeester & ten Dijke J Pathol 2011, 223: 205-218). Mutations of TGFβ signalling components such as receptors and Smad proteins decrease the tumour-suppressive function of this growth factor (Meulmeester & ten Dijke J Pathol 2011, 223: 205-218). TGFβ induces angiogenesis even though its role is more vast and complex (Massague, J. Cell 2008, 134: 215-230). However, we have shown that a close relative, ActivinA, is anti-angiogenic in vivo (Panopoulou et al., Cancer Res 2005: 65, 1877-1886). Thus, the research plans (RP) concerning TGFβ signalling are described below.

Trafficking and signalling machinery of ActivinA

Rationale: TGFβ receptors localise to both raft and non-raft membrane domains and the internalisation route dictates whether signalling or degradation will ensue. Internalisation of TGFβ receptors, via the clathrin-coated pathway into an EEA1 and SARA (SMAD Anchor for Receptor Activation) positive endosomes, where SARA recruits non-phosphorylated SMAD2/3 to the activated receptors for phosphorylation, and promotes downstream signalling (Tsukazaki et al. Cell, 1998, 95: 779-791). However, internalisation via the raft-caveolar pathway, where SMAD7 and SMURF2 are localised, promotes ubiquitin-dependent receptor degradation. Indeed, inhibition of this pathway leads to receptor stabilisation, suggesting that trafficking of receptors to the SARA positive early endosomes (Panopoulou et al. J. Biol. Chem. 2002, 277: 18046-52) functions to sequester receptors from rafts and caveolae, thereby stabilising the receptors (Di Guglielmo et al., Nat Cell Biol. 2003, 5: 410-21), (Felberbaum-Corti et al, Nat Cell Biol. 2003, 5: 382). Thus, partitioning between these two internalisation pathways appears to be a dynamic and balanced process influencing the signalling outcome of the activated TGFβ family receptors.

However, as mentioned above, in addition to clathrin-mediated and caveolar routes ligands /receptor complexes can be internalised via macropinocytosis, the APPL pathway, or the non-clathrin and non-caveolar pathway that ferries lipid raft components and extracellular fluid into the GEEC (GPI-anchored protein enriched early endosomal compartment). There is limited information concerning internalisation of the TGFβ receptors complexes via any of the alternative routes and the consequences on signalling. Even less is known about the internalisation of ActivinA/receptor complexes. 

Approach-Aims: Based on the above, it is evident that there is vast lack of knowledge about the internalisation routes of ActivinA/receptor complexes. Also, SARA, an immediate downstream component of the TGFβ/ActivinA signalling cascade, has very interesting properties that justify further attention to this protein. Therefore our approach is

  • To define the trafficking routes of various GFR complexes in cultured cells with an unprecedented degree of precision, combining confocal microscopy and automated image analysis.
    Using ActivinA labelled with Alexa 488 (in collaboration with Marco Hyvonen, Cambridge University, UK), tagged receptors and specific markers of various endocytic compartments we are investigating the trafficking route of ActivinA and its receptors. This work is ongoing and has defined the mechanism of ActivinA internalisation and it’s role in signalling.
  • To define the molecular machinery responsible for this transport, using proteomics and functional genomics approaches
    Identification of the molecular machinery is underway. SARA is a FYVE domain containing protein which interacts with SMAD proteins and localises to the early endocytic compartment. We have identified 10 SARA-interactive proteins using a two-hybrid screen. Investigation of two of them is already completed and published. The one is ERBIN (Sflomos, et al., J Cell Sci 2011: 124, 3209-3222) and the other is RNF11 (Kostaras et al., Oncogene 32 (2013) 5220-32.). We have also identified Alk4 and ActrIIB-interacting proteins using yeast 2 hybrid screens and proteomics and analysis of their role in trafficking/signalling is being investigated.
  • To define the signalling networks downstream of receptor activation
    In collaboration with the High Throughput Facility at the Max Planck Institute in Dresden we have carried out a phosphatase and kinase siRNA screen which has revealed the fingerprint of the ActivinA signalling network. Bioinformatics has been performed and a secondary validation screen is partially completed. We will now focus on regulatory nodes which control ActivinA signalling.
  • To integrate the trafficking/signalling interconnection in disease models
    We have recently initiated a project involving the role of altered trafficking in a rare disease model of Fibrodysplasia Ossificans Progressiva (FOP) characterised by mutations of Alk2, the type 1 BMP receptor.
  • To investigate ligand/receptor signalling/trafficking circuitries in hESCs and hiPSCs and elucidate the alterations that occur during differentiation in these circuitries and the mechanism thereof (with emphasis on differentiation towards mesoderm).
    We are comparing the trafficking routes in human stem cells and induced pluripotent stem cells to those of the same cell types induced to differentiate along the endothelial cell lineage. Initial findings indicate altered trafficking pathways as the cell exits the pluripotent state and enters a differentiation program.

Interconnection between membrane trafficking and the centrosome cycle

The centrosome is the major microtubule-organizing center of animal cells, which influences cell shape and directs the formation of the bipolar mitotic spindle (Nigg and Stearns, Nat Cell Biol., 2011, 13: 1154-60). At the core of a typical centrosome are two cylindrical microtubule-based structures termed centrioles, which recruit a matrix of associated pericentriolar material. Aberrations in numerical and structural integrity of centrosomes interfere with spindle formation and chromosome segregation. Furthermore, the centrosome plays a role in asymmetric cell division (Rusan and Peifer, J. Cell Biol., 2007: 13-20).

Centrosome positioning is crucial, as it determines the location of many other organelles within the cell and regulates proper cell division. This localization is actively maintained via interconnected mechanical forces applied from microtubules and the cortical actin network. This crosstalk is essential not only for the maintenance of the central localization of the MTOC in interphase cells, but also for orchestrating the movement of the duplicated centrosomes during cell division. Changes in numbers, structure and/or location of centrosomes result in phenotypic abnormalities (from apoptosis to uncontrolled proliferation) due to chromosome instabilities (Ganem et al., Nature, 2009, 460: 278-82, Quintyne et al., Science, 2005, 307: 127-9). Centrosome amplification forces cells to pass through a multipolar spindle intermediate, enhancing merotelic attachments and lagging chromosomes, and as a consequence unequal chromosome segregation into the daughter cells. In fact, altered centrosomes have been described in many tumors. Furthermore, direct manipulation of centrosomes in vitro leads to aneuploidy and transformation, supporting a potentially causative effect towards malignancy (Zhou, et al., Nat Genet., 1998, 20: 189-93).

Role of RhoD in the centrosome cycle, cell division and cancer

Rationale: Some years ago we identified a novel Rho protein localizing to the early endocytic compartment (RhoD). As our main area of interest is trafficking and signalling in this compartment we investigated the role of RhoD and found that it controlled early endosome vesicular motility (Murphy et al., Nature 1996, 384:427-432). We also found an involvement of RhoD in centrosome duplication and possible links between the centrosome’s structural and functional integrity to vesicular trafficking (Kyrkou et al., Oncogene 2013, 32:1831-1842 and Kyrkou et al., Small GTPases 2013, 4). The role of vesicular trafficking in the regulation of the centrosome cycle has been largely unexplored. Recently, however, several studies have indicated the involvement of molecules and/or complexes of the trafficking routes in centrosome positioning, duplication and regulation. Functional screens have revealed communication between the outer nuclear envelope, the Golgi apparatus, the endosomal recycling compartment and centrosomes, while other studies underline the involvement of the ESCRT complex proteins in centrosome function. In the transgenic mice we generated expressing RhoD we observed supra-basal cell proliferation, which is suggestive of alterations in the balance between symmetric/asymmetric cell divisions in the epidermis (Kyrkou et al., Oncogene 2013, 32:1831-1842 and Kyrkou et al., Small GTPases 2013, 4).

We are further investigating the role of RhoD in centrosome function and organization and cell division.

Other activities

Publications

Dimitris Basagiannis, Sofia Zografou, Carol Murphy, Theodore Fotsis, Lucia Morbidelli, Marina Ziche, Christopher Bleck, Jason Mercer, and Savvas Christoforidis. VEGF induces signalling and angiogenesis by directing VEGFR2 internalisation via macropinocytosis. J. Cell Sci  2016 in press.

Tsolis K, Bagli E, Kanaki K, Zografou S, Carpentier S, Bei E, Savvas Christoforidis S,  Zervakis M, Murphy C, Fotsis T, Economou A. Proteome changes during transition from human embryonic to vascular progenitor cells. Accepted in J Proteome Research, 2016, 15: 1995-2007.

Kyrkou A, Stellas D, Syrrou M, Klinakis A, Fotsis T, Murphy C: Generation of human induced pluripotent stem cells in defined, feeder-free conditions. Lab Resource, Stem Cell Research, 2016 in press. http://www.sciencedirect.com/science/article/pii/S1873506116300381

Kouroupis D, Kyrkou A, Triantafyllidi E, Katsimpoulas M, Chalepakis G, Goussia A,  Georgoulis A, Murphy C, Fotsis T: Generation of stem cell-based bioartificial anterior cruciate ligament (ACL) grafts for effective ACL rupture repair. Stem Cell Research, 2016 in press. http://www.ncbi.nlm.nih.gov/pubmed/27217303

Karali E, Bellou S, Stellas Dimitris, Klinakis Apostolos Murphy C, Fotsis T: ER mediates induction of endothelial cell survival and angiogenesis by VEGF: PLCg via mTORC1 activates ATF6 and PERK. Mol. Cell 2014, 54: 559-72.

Kostaras E, Pedersen NM, Stenmark H, Fotsis T, Murphy C: SARA and RNF11 at the crossroads of EGFR signalling and trafficking. Methods Enzymol 2014, 535: 225-47.

Bellou S, Pentheroudakis G, Murphy C, Fotsis T: Anti-angiogenesis in cancer therapy: Hercules and Hydra. Cancer Lett. 2013, 338: 291-28.

Kyrkou A, Soufi M, Bahtz R, Ferguson C, Bai M, Parton RG, Hoffmann I, Zerial M, Fotsis T, Murphy C: RhoD participates in the regulation of cell-cycle progression and centrosome duplication. Oncogene 2013, 32:1831-1842.

Kyrkou A, Soufi M, Bahtz R, Ferguson C, Bai M, Parton RG, Hoffmann I, Zerial M, Fotsis T, Murphy C: The RhoD to centrosomal duplication. Small GTPases 2013, 4: 116-22.

Kostaras E, Sflomos G, Pedersen NM, Stenmark H, Fotsis T*, Murphy C*: SARA and RNF11 interact with each other and ESCRT-0 core proteins and regulate degradative EGFR trafficking. Oncogene 2013, 32: 5220-32. * joint last authors.

Bellou S, Karali E, Bagli E, Al-Maharik N, Morbidelli L, Ziche M, Adlercreutz H, Murphy C, Fotsis T: The isoflavone metabolite 6-methoxyequol inhibits angiogenesis and suppresses tumor growth. Mol Cancer 2012, 11:35.

Sflomos G, Kostaras E, Panopoulou E, Pappas N, Kyrkou A, Politou AS, Fotsis T, Murphy C: ERBIN is a new SARA-interacting protein: competition between SARA and SMAD2 and SMAD3 for binding to ERBIN. J Cell Sci 2011, 124:3209-3222.

Bellou S, Hink MA, Bagli E, Panopoulou E, Bastiaens PI, Murphy C, Fotsis T: VEGF autoregulates its proliferative and migratory ERK1/2 and p38 cascades by enhancing the expression of DUSP1 and DUSP5 phosphatases in endothelial cells. Am J Physiol Cell Physiol 2009, 297:C1477-1489.

Kardassis D, Murphy C, Fotsis T, Moustakas A, Stournaras C: Control of transforming growth factor beta signal transduction by small GTPases. FEBS J 2009, 276:2947-2965.

Murphy C: Endo-fin-ally a SARA for BMP receptors. J Cell Sci 2007, 120:1153-1155.

Fyhrquist P, Mwasumbi L, Vuorela P, Vuorela H, Hiltunen R, Murphy C, Adlercreutz H: Preliminary antiproliferative effects of some species of Terminalia, Combretum and Pteleopsis collected in Tanzania on some human cancer cell lines. Fitoterapia 2006, 77:358-366.

Papanikolaou A, Papafotika A, Murphy C, Papamarcaki T, Tsolas O, Drab M, Kurzchalia TV, Kasper M, Christoforidis S: Cholesterol-dependent lipid assemblies regulate the activity of the ecto-nucleotidase CD39. J Biol Chem 2005, 280:26406-26414.

Panopoulou E, Murphy C, Rasmussen H, Bagli E, Rofstad EK, Fotsis T: Activin A suppresses neuroblastoma xenograft tumor growth via antimitotic and antiangiogenic mechanisms. Cancer Res 2005, 65:1877-1886.

Bagli E, Stefaniotou M, Morbidelli L, Ziche M, Psillas K, Murphy C, Fotsis T: Luteolin inhibits vascular endothelial growth factor-induced angiogenesis; inhibition of endothelial cell survival and proliferation by targeting phosphatidylinositol 3'-kinase activity. Cancer Res 2004, 64:7936-7946.

Panopoulou E, Gillooly DJ, Wrana JL, Zerial M, Stenmark H, Murphy C*, Fotsis T*: Early endosomal regulation of Smad-dependent signalling in endothelial cells. J Biol Chem 2002, 277:18046-18052.* joint last authors.

Karetsou Z, Kretsovali A, Murphy C, Tsolas O, Papamarcaki T: Prothymosin alpha interacts with the CREB-binding protein and potentiates transcription. EMBO Rep 2002, 3:361-366.

Hatzi E, Murphy C, Zoephel A, Rasmussen H, Morbidelli L, Ahorn H, Kunisada K, Tontsch U, Klenk M, Yamauchi-Takihara K, et al.: N-myc oncogene overexpression down-regulates IL-6; evidence that IL-6 inhibits angiogenesis and suppresses neuroblastoma tumor growth. Oncogene 2002, 21:3552-3561.

Hatzi E, Murphy C, Zoephel A, Ahorn H, Tontsch U, Bamberger AM, Yamauchi-Takihara K, Schweigerer L, Fotsis T: N-myc oncogene overexpression down-regulates leukemia inhibitory factor in neuroblastoma. Eur J Biochem 2002, 269:3732-3741.

Murphy C, Saffrich R, Olivo-Marin JC, Giner A, Ansorge W, Fotsis T, Zerial M: Dual function of rhoD in vesicular movement and cell motility. Eur J Cell Biol 2001, 80:391-398.

Hatzi E, Breit S, Zoephel A, Ashman K, Tontsch U, Ahorn H, Murphy C, Schweigerer L, Fotsis T: MYCN oncogene and angiogenesis: down-regulation of endothelial growth inhibitors in human neuroblastoma cells. Purification, structural, and functional characterization. Adv Exp Med Biol 2000, 476:239-248.

McBride HM, Rybin V, Murphy C, Giner A, Teasdale R, Zerial M: Oligomeric complexes link Rab5 effectors with NSF and drive membrane fusion via interactions between EEA1 and syntaxin 13. Cell 1999, 98:377-386.

Zacchi P, Stenmark H, Parton RG, Orioli D, Lim F, Giner A, Mellman I, Zerial M, Murphy C: Rab17 regulates membrane trafficking through apical recycling endosomes in polarized epithelial cells. J Cell Biol 1998, 140:1039-1053.

Simonsen A, Lippe R, Christoforidis S, Gaullier JM, Brech A, Callaghan J, Toh BH, Murphy C, Zerial M, Stenmark H: EEA1 links PI(3)K function to Rab5 regulation of endosome fusion. Nature 1998, 394:494-498.

Murphy C, Zacchi P, Parton RG, Zerial M, Lim F: HSV infection of polarized epithelial cells on filter supports: implications for transport assays and protein localization. Eur J Cell Biol 1997, 72:278-281.

Zuniga Mejia Borja A, Murphy C, Zeller R: AltFGF-2, a novel ER-associated FGF-2 protein isoform: its embryonic distribution and functional analysis during neural tube development. Dev Biol 1996, 180:680-692.

Murphy C, Saffrich R, Grummt M, Gournier H, Rybin V, Rubino M, Auvinen P, Lutcke A, Parton RG, Zerial M: Endosome dynamics regulated by a Rho protein. Nature 1996, 384:427-432.

Kretschmer C, Murphy C, Biesinger B, Beckers J, Fickenscher H, Kirchner T, Fleckenstein B, Ruther U: A Herpes saimiri oncogene causing peripheral T-cell lymphoma in transgenic mice. Oncogene 1996, 12:1609-1616.

Murphy C, Zerial M: Expression of Rab proteins during mouse embryonic development. Methods Enzymol 1995, 257:324-332.

Murphy C, Beckers J, Ruther U: Regulation of the human C-reactive protein gene in transgenic mice. J Biol Chem 1995, 270:704-708.

Murphy C, Kretschmer C, Biesinger B, Beckers J, Jung J, Desrosiers RC, Muller-Hermelink HK, Fleckenstein BW, Ruther U: Epithelial tumours induced by a herpesvirus oncogene in transgenic mice. Oncogene 1994, 9:221-226.

Lutcke A, Parton RG, Murphy C, Olkkonen VM, Dupree P, Valencia A, Simons K, Zerial M: Cloning and subcellular localization of novel rab proteins reveals polarized and cell type-specific expression. J Cell Sci 1994, 107 (Pt 12):3437-3448.

Fotsis T, Murphy C, Gannon F: Nucleotide sequence of the bovine insulin-like growth factor 1 (IGF-1) and its IGF-1A precursor. Nucleic Acids Res 1990, 18:676.

Murphy C, Fotsis T, Pantzar P, Adlercreutz H, Martin F: Analysis of tamoxifen and its metabolites in human plasma by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring (SIM). J Steroid Biochem Mol Biol 1987, 26:547-555.

Murphy C, Fotsis T, Pantzar P, Adlercreutz H, Martin F: Analysis of tamoxifen, N-desmethyltamoxifen and 4-hydroxytamoxifen levels in cytosol and KCl-nuclear extracts of breast tumours from tamoxifen treated patients by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring (SIM). J Steroid Biochem Mol Biol 1987, 28:609-618.

Murphy C, Fotsis T, Adlercreutz H, Martin F: Analysis of tamoxifen and 4-hydroxytamoxifen levels in immature rat uterine cytoplasm and KCl-nuclear extracts by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring (SIM). J Steroid Biochem Mol Biol 1987, 28:289-299.