A role for exosomes in the constitutive and stimulus-induced ectodomain cleavage of L1 and CD44.
Ectodomain shedding is a proteolytic mechanism by which transmembrane molecules are converted into a soluble form. Cleavage is mediated by metalloproteases and proceeds in a constitutive or inducible fashion. Although believed to be a cell-surface event, there is increasing evidence that cleavage can take place in intracellular compartments. However, it is unknown how cleaved soluble molecules get access to the extracellular space. By analysing L1 (CD171) and CD44 in ovarian carcinoma cells, we show in the present paper that the cleavage induced by ionomycin, APMA (4-aminophenylmercuric acetate) or MCD (methyl-beta-cyclodextrin) is initiated in an endosomal compartment that is subsequently released in the form of exosomes. Calcium influx augmented the release of exosomes containing functionally active forms of ADAM10 (a disintegrin and metalloprotease 10) and ADAM17 [TACE (tumour necrosis factor a-converting enzyme)] as well as CD44 and L1 cytoplasmic cleavage fragments. Cleavage could also proceed in released exosomes, but only depletion of ADAM10 by small interfering RNA blocked cleavage under constitutive and induced conditions. In contrast, cleavage of L1 in response to PMA occurred at the cell surface and was mediated by ADAM17. We conclude that different ADAMs are involved in distinct cellular compartments and that ADAM10 is responsible for shedding in vesicles. Our findings open up the possibility that exosomes serve as a platform for ectodomain shedding and as a vehicle for the cellular export of soluble molecules.
Stoeck
Alexander
A
Tumor Immunology Programme, D010, German Cancer Research Center, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany.
Keller
Sascha
S
Riedle
Svenja
S
Sanderson
Michael P
MP
Runz
Steffen
S
Le Naour
Francois
F
Gutwein
Paul
P
Ludwig
Andreas
A
Rubinstein
Eric
E
Altevogt
Peter
P
D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism; D000072197:ADAM10 Protein; D000072198:ADAM17 Protein; D053829:Amyloid Precursor Protein Secretases; D045744:Cell Line, Tumor; D022162:Cytoplasmic Vesicles / Q000378:metabolism*; D005089:Exocytosis; D005786:Gene Expression Regulation; D006801:Humans; D018960:Hyaluronan Receptors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D039842:Neural Cell Adhesion Molecule L1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D034622:RNA Interference
D018960:Hyaluronan Receptors; D008565:Membrane Proteins; D039842:Neural Cell Adhesion Molecule L1; D053829:Amyloid Precursor Protein Secretases; D051722:ADAM Proteins; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human