Activated leukocyte cell adhesion molecule (ALCAM) regulation of tumor cell behavior and neuronal targeting

..........................................................................................................59 Introduction .....................................................................................................60 Results .............................................................................................................63 ALCAM expression correlates with cell motility in uveal melanoma cell lines .................................................................................63 Establishment of ALCAM-silenced MUM-2B cell lines by shRNA knockdown ..............................................................................................65 ALCAM-silenced cells display reduced motility and invasive capacity ....................................................................................................66 ALCAM overexpression is not sufficient to enhance migration and invasive capacity in MUM-2C cells ........................................................67 ALCAM-silenced cells exhibit reduced activation of MMP-2 ...............67 Cadherin-based junctions are disorganized in ALCAM-silenced cells ..........................................................................................................68 ALCAM expression enhances cadherin-mediated cell-cell contacts in the 2C-ALC cell line ...........................................................................70 Discussion .......................................................................................................70 CHAPTER FOUR: IDENTIFICATION OF NOVEL ALCAM EXTRAAND INTRACELLULAR LIGANDS .....................................................................88 Abstract ...........................................................................................................88 Introduction .....................................................................................................89 L1 cell adhesion molecule, an ALCAM extracellular interactor ............89 Intracellular interactors ............................................................................91 Results .............................................................................................................91 L1 and ALCAM are expressed in the developing visual pathway ..........91 Evidence for a trans-interaction between ALCAM and L1 .....................95 ALCAM and L1 interact in trans to regulate retinotopic mapping .........96 Isolation of eight candidate intracellular interactors by affinity chromatography .......................................................................................97 IQGAP1 ...................................................................................................97 Filamin A .................................................................................................98 α-actinin-4 ...............................................................................................99 Final comments .......................................................................................99 CHAPTER FIVE: CONLCUSIONS AND FUTURE DIRECTIONS ............................106..........................................................................................................88 Introduction .....................................................................................................89 L1 cell adhesion molecule, an ALCAM extracellular interactor ............89 Intracellular interactors ............................................................................91 Results .............................................................................................................91 L1 and ALCAM are expressed in the developing visual pathway ..........91 Evidence for a trans-interaction between ALCAM and L1 .....................95 ALCAM and L1 interact in trans to regulate retinotopic mapping .........96 Isolation of eight candidate intracellular interactors by affinity chromatography .......................................................................................97 IQGAP1 ...................................................................................................97 Filamin A .................................................................................................98 α-actinin-4 ...............................................................................................99 Final comments .......................................................................................99 CHAPTER FIVE: CONLCUSIONS AND FUTURE DIRECTIONS ............................106 The role of ALCAM in regulating N-cadherin-based adherens junctions ...107 IQGAP1 regulation of adherens junctions ............................................110 Evidence for IQGAP1/ALCAM crosstalk .............................................113 Filamin A regulation of transmembrane proteins at the cell surface .....117 Evidence for crosstalk between filamin A and ALCAM ......................118 α-actinin-4 regulation of the cadherin-catenin axis ...............................120