Umeå University Medical Dissertations New Series No 1080 • ISSN 0346-6612 • ISBN 978-91-7264-246-1 ___________________________________________________________________________ Protein Tyrosine Kinases and The Regulation of Signalling and Adhesion in Drosophila melanogaster Caroline Grabbe Umeå Centre for Molecular Pathogenesis Umeå University Umeå, Sweden, 2007 Copyright © 2007 by Caroline Grabbe ISBN 978-91-7264-246-1 _______________________________________________________ Printed in Sweden by Solfjädern Offset AB Umeå, 2007 2 To Chance, and Life Itself… (I det långa loppet träffar man bara det man siktat på. Så även om man missar sitt mål i första försöket så är det lika bra att sikta högt.) Henry David Thoreau 3 TABLE OF CONTENTS ABBREVIATIONS ................................................................................................................ 6 SUMMARY............................................................................................................................. 8 PAPERS INCLUDED IN THIS THESIS............................................................................. 9 PAPERS NOT INCLUDED IN THIS THESIS ................................................................... 9 INTRODUCTION................................................................................................................ 10 Drosophila melanogaster as a model system....................................................................... 10 Historical perspective............................................................................................................. 10 Why Drosophila? ................................................................................................................... 11 Drosophila genetic tools and techniques................................................................................ 11 Drosophila life cycle and development.................................................................................. 13 Development and attachment of Drosophila muscles........................................................ 13 Myoblast fusion...................................................................................................................... 14 Duf/Kirre, Sticks and stones ............................................................................................ 15 Org-1, Jelly belly ............................................................................................................. 16 Drosophila Neuromuscular Junctions (NMJs)....................................................................... 17 Cell adhesion......................................................................................................................... 18 Cell-matrix adhesions in general and focal adhesions in particular ....................................... 18 Extracellular matrix................................................................................................................ 19 Integrins.................................................................................................................................. 19 Integrins in Drosophila .......................................................................................................... 21 Integrin-associated proteins.................................................................................................... 22 Actin ................................................................................................................................ 22 Src-family kinases (SFKs) ............................................................................................... 22 p130Cas ........................................................................................................................... 23 Talin................................................................................................................................. 23 Paxillin............................................................................................................................. 24 The PINCH-ILK-Parvin (PIP) complex .......................................................................... 24 α-actinin........................................................................................................................... 24 Vinculin ........................................................................................................................... 25 Tensin .............................................................................................................................. 25 Tiggrin ............................................................................................................................. 25 Cell-Cell Adhesion – The AJC complex ............................................................................. 25 Tight Junctions (TJs) ....................................................................................................... 26 Adherens Junctions (AJs) ................................................................................................ 26 Cell Signalling....................................................................................................................... 27 Protein Kinases and Phosphorylation..................................................................................... 27 Focal Adhesion Kinase......................................................................................................... 28 Structural features of FAK ..................................................................................................... 29 The N-terminal FERM domain .............................................................................................. 29 The catalytic kinase domain................................................................................................... 30 The Focal Adhesion Targeting (FAT) domain....................................................................... 30 Additional conserved motifs in FAK ..................................................................................... 31 FRNK (FAK-related non-kinase)........................................................................................... 31 FAK function ........................................................................................................................ 32 Regulation of FAK activity .................................................................................................... 32 FAK and adhesion.................................................................................................................. 33 FAK and migration................................................................................................................. 34 FAK in tumour development and progression ....................................................................... 35 The role of FAK in mammalian systems in vivo.................................................................... 36 FAK – A bridge between Integrins and RTKs? ..................................................................... 37 Pyk2/RAFTK/CAKβ............................................................................................................ 38 FAK and Pyk2 interrelationship; Redundant and specific functions?.................................... 39 4 FAK and Pyk2 in the nervous system .................................................................................... 39 Drosophila FAK – Fak56 ..................................................................................................... 41 Characterisation of the Fak56 gene........................................................................................ 41 Physiological functions of Fak56 in the fruit fly.................................................................... 42 Bang Sensitivity .................................................................................................................... 43 The Giant Fiber pathway........................................................................................................ 44 Electrophysiology................................................................................................................... 45 Bang Sensitivity in vertebrates? ............................................................................................. 46 Regulation of Cellular Signalling........................................................................................ 47 Receptor Tyrosine Kinases (RTKs)........................................................................................ 47 The ERK/MAPK signalling module ...................................................................................... 47 Anaplastic Lymphoma Kinase ............................................................................................ 48 Structural Features of ALK .................................................................................................... 49 Signalling mediated via the ALK receptor............................................................................. 50 ALK ligands..................................................................................................................... 50 The PLC-γ pathway ......................................................................................................... 50 The RAS/MAPK pathway ............................................................................................... 50 The PI3K pathway ..........................................................................................................