Sethi, Tariq

Elliott, Paul Anthony

2010-11-22T14:05:47Z

2010-11-22T14:05:47Z

2008

Integrins are heterodimeric transmembrane proteins that provide a bi-directional link between the cell’s internal biological mechanisms and the extracellular environment. During inside-out signalling, intracellular messages converge on the integrin cytoplasmic domain to induce a conformational change. This is transmitted to the extracellular domain where it results in an alteration in affinity for integrin ligands such as fibronectin and laminin. In this way the cell has developed the ability to modulate the critical functions of adhesion and cell movement. In outside-in signalling, the integrin performs a more complex function than simple adhesion; upon binding to ligand, the integrin extracellular domain undergoes a conformational change which is transmitted to the cytoplasmic domain. This alters the integrin’s cytoplasmic domain affinity for intracellular signalling proteins and results in the activation of intracellular second messenger pathways. In this way, the extracellular milieu is able to influence intracellular signalling including those involved in apoptosis. This thesis demonstrates data which provide original insights into bi-directional integrin signalling: Inside-out signalling: Constitutively active Notch1 increases β3-integrin affinity and abrogates Hras-mediated integrin suppression without increasing expression of β3- integrin. Dominant-Negative Rras blocks Notch-mediated integrin activation and Notch1-mediated reversal of Hras and Raf-mediated integrin suppression and this is independent of erk phosphorylation. Notch1 induces Rras activation. Functional adhesion assays confirm that Notch1IC increases K562 adhesion in a β1-integrin dependent manner and this is abrogated by Dominant-Negative Rras. This data supports a mechanism in which Notch1 increases integrin affinity via activation of Rras. Outside-in signalling: Evidence is presented demonstrating that extracellular matrix proteins, laminin and fibronectin, activate β1-integrins to protect SCLC cells against the apoptotic effects of etoposide and ionizing radiation via PI3Kinase activation. This occurs in two ways: 1) PI3Kinase-dependent β1-integrin signalling resulting in phosphorylation of Bad and reduced caspase-9 cleavage and 2) a β1-integrinmediated over-riding of etoposide and radiotherapy-induced cell cycle S phase delay and G2/M arrest. β1-integrin-mediated outside-in survival signalling was investigated further in the in vivo setting; MatrigelTM, a basement membrane product rich in extracellular matrix proteins, promoted SCLC xenograft survival and growth in a β1-integrin and tyrosine kinase-dependent manner. This data provides novel insights into the critical functions that integrins play in adhesion and survival signalling.

http://hdl.handle.net/1842/4393

en

The University of Edinburgh

PS Hodkinson, T Elliott, WS Wong, RC Rintoul, AC Mackinnon, C Haslett and T Sethi, "ECM overrides DNA damage-induced cell cycle arrest and apoptosis in small-cell lung cancer cells through b1 integrin-dependent activation of PI3-kinase", Cell Death and Differentiation (2006) 13, 1776–1788,

Philip S. Hodkinson, Paul. A. Elliott, Yatish Lad, Brian J. McHugh, Alison C MacKinnon Christopher Haslett and Tariq Sethi, "Mammalian notch-1 activates β1 integrins via the small gtpase r-ras." JBC Papers in Press. Published on July 30, 2007 as Manuscript M703601200 The latest version is at http://www.jbc.org/cgi/doi/10.1074/jbc.M703601200

integrin

signalling

apoptosis

Integrin affinity modulation and survival signalling.

Thesis or Dissertation

Doctoral

PhD Doctor of Philosophy