LAMP proteins are required for fusion of lysosomes with phagosomes.
Lysosome-associated membrane proteins 1 and 2 (LAMP-1 and LAMP-2) are delivered to phagosomes during the maturation process. We used cells from LAMP-deficient mice to analyze the role of these proteins in phagosome maturation. Macrophages from LAMP-1- or LAMP-2-deficient mice displayed normal fusion of lysosomes with phagosomes. Because ablation of both the lamp-1 and lamp-2 genes yields an embryonic-lethal phenotype, we were unable to study macrophages from double knockouts. Instead, we reconstituted phagocytosis in murine embryonic fibroblasts (MEFs) by transfection of FcgammaIIA receptors. Phagosomes formed by FcgammaIIA-transfected MEFs obtained from LAMP-1- or LAMP-2- deficient mice acquired lysosomal markers. Remarkably, although FcgammaIIA-transfected MEFs from double-deficient mice ingested particles normally, phagosomal maturation was arrested. LAMP-1 and LAMP-2 double-deficient phagosomes acquired Rab5 and accumulated phosphatidylinositol 3-phosphate, but failed to recruit Rab7 and did not fuse with lysosomes. We attribute the deficiency to impaired organellar motility along microtubules. Time-lapse cinematography revealed that late endosomes/lysosomes as well as phagosomes lacking LAMP-1 and LAMP-2 had reduced ability to move toward the microtubule-organizing center, likely precluding their interaction with each other.
Huynh
Kassidy K
KK
Division of Cell Biology, The Hospital for Sick Children, Toronto, Canada.
Eskelinen
Eeva-Liisa
EL
Scott
Cameron C
CC
Malevanets
Anatoly
A
Saftig
Paul
P
Grinstein
Sergio
S
D000818:Animals; D002478:Cells, Cultured; D052119:Lysosomal-Associated Membrane Protein 2 / Q000235:genetics / Q000502:physiology; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000502:physiology*; D008247:Lysosomes / Q000378:metabolism / Q000502:physiology*; D008264:Macrophages / Q000502:physiology / Q000648:ultrastructure; D008561:Membrane Fusion; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D010587:Phagocytosis / Q000235:genetics; D010588:Phagosomes / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins; D052118:Lysosomal-Associated Membrane Protein 1
D052119:Lysosomal-Associated Membrane Protein 2; D051907:Lysosomal Membrane Proteins; D020691:rab GTP-Binding Proteins; D000090802:rab7 GTP-Binding Proteins; C052255:Lamp1 protein, mouse; D052118:Lysosomal-Associated Membrane Protein 1; C000716465:rab7 GTP-binding proteins, mouse