Receptor-mediated uptake of an extracellular Bcl-x(L) fusion protein inhibits apoptosis.
Bcl-x(L), a member of the Bcl-2 family, inhibits many pathways of apoptosis when overexpressed in the cell cytosol. We examined the capacity of Bcl-x(L) fusion proteins to bind cells from the outside and block apoptosis. Full-length Bcl-x(L) protein at micromolar concentrations did not affect apoptosis when added to cell media. To increase uptake by cells, Bcl-x(L) was fused to the receptor-binding domain of diphtheria toxin (DTR). The Bcl-x(L)-DTR fusion protein blocked apoptosis induced by staurosporine, gamma-irradiation, and poliovirus in a variety of cell types when added to media. The potency of inhibition of poliovirus-induced apoptosis by Bcl-x(L)-DTR was greater than that of strong caspase inhibitors. Brefeldin A, an inhibitor of vesicular traffic between the endoplasmic reticulum and Golgi apparatus, prevented the Bcl-x(L)-DTR blockade of apoptosis induced by staurosporine, suggesting that Bcl-x(L)-DTR must be endocytosed and reach intracellular compartments for activity. Many diseases are caused by overexpression or underexpression of Bcl-x(L) homologues. Extracellular delivery of Bcl-2 family member proteins may have a wide range of uses in promoting or preventing cell death.
Liu
X H
XH
Biochemistry Section, Surgical Neurology Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA.
Castelli
J C
JC
Youle
R J
RJ
D017209:Apoptosis / Q000187:drug effects*; D020126:Brefeldin A / Q000494:pharmacology; D004167:Diphtheria Toxin / Q000235:genetics / Q000494:pharmacology*; D004705:Endocytosis; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011487:Protein Conformation; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology; D051020:bcl-X Protein
C494811:BCL2L1 protein, human; D004167:Diphtheria Toxin; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D051020:bcl-X Protein; D020126:Brefeldin A; D011065:Poly(ADP-ribose) Polymerases; D019311:Staurosporine