title: Development of a bidirectional caspase-3 expression system for the induction of apoptosis.
abstract: Caspase-3 is the executioner caspase of apoptosis whose activation in mammalian cells represents the last stage of the programmed cell death signaling pathway and the initiation of the lethal digestion of cell proteins. Active caspase-3 is a tetramer composed of two p12 and two p17 subunits derived from cleavage of procaspase-3 during activation. Here, we armed GFP-fusion proteins of both the caspase-3 p12 and p17 subunits with signals from Ig-kappa light chain that allows its efficient secretion from the cells (Sec) and from HIV-1 Tat that facilitates its uptake and nuclear translocation by other cells (NLS). We found that treatment of cells with conditioned media from cells expressing both Sec-GFP-p17-NLS and Sec-GFP-p12-NLS was able to transduce active caspase-3 with consequent cell death of treated cultures. Use of various combinations of constructs demonstrated that both subunits were required and that each one needed to possess both Sec and NLS. Our observations introduce a bidirectional protein transduction system with the ability to introduce active caspase-3 into cells and cause apoptosis. This system may have important therapeutic applications.
authors: Department of Neuroscience, Center for Neurovirology, Temple University School of Medicine, Philadelphia, Pennsylvania 19122, USA.
authors: Martyn K
authors: MK
authors: White
authors: Shohreh
authors: S
authors: Amini
authors: Kamel
authors: K
authors: Khalili
authors: Michael
authors: M
authors: Kogan
authors: Keri
authors: K
authors: Donaldson
authors: Nune
authors: N
authors: Darbinian
mesh_terms: D021581:Active Transport, Cell Nucleus; D017209:Apoptosis; D001483:Base Sequence; D053148:Caspase 3 / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D008969:Molecular Sequence Data; D013347:Subcellular Fractions
chemical_list: D049452:Green Fluorescent Proteins; D053148:Caspase 3